Biotechnology: Principles and Processes

Topic: RNA Interference and Transgenic Organisms; Subtopic: Host-mediated RNAi for Nematode Resistance

Keyword Definitions:

• Nematode: A parasitic roundworm affecting plants or animals, causing diseases or crop losses.

• Transgenic Host: An organism that has incorporated foreign genes into its genome through genetic engineering.

• RNA Interference (RNAi): A biological process where double-stranded RNA triggers degradation of complementary mRNA, silencing specific gene expression.

• Interfering RNA: RNA molecules (siRNA or miRNA) that mediate RNAi to regulate gene expression or defend against pathogens.

• Sense and Antisense RNA: Complementary RNA strands; pairing leads to formation of double-stranded RNA essential for RNAi.

• Gene Silencing: Process of turning off the expression of a specific gene at the transcriptional or post-transcriptional level.

• Host-mediated Resistance: Resistance conferred by introducing pathogen-specific genes into a host organism to trigger defense mechanisms.

Lead Question – 2022 (Abroad)
Given below are two statements: one is labelled as Assertion (A) and the other is labelled as Reason (R).
Assertion (A): The nematode cannot survive in a transgenic host which expresses specific interfering RNA.
Reason (R): Nematode-specific gene introduced in the host produces both sense and antisense complementary RNA which initiate RNA interference in the host cell.
In the light of the above statement, choose the most appropriate answer from the options given below:
1. (A) is correct but (R) is not correct
2. (A) is not correct but (R) is correct
3. Both (A) and (R) are correct and (R) is the correct explanation of (A)
4. Both (A) and (R) are correct but (R) is not the correct explanation of (A)

Explanation:
Correct answer is option 3. In host-mediated RNA interference, a transgenic plant expresses nematode-specific double-stranded RNA. The host produces both sense and antisense RNA, forming double-stranded RNA that triggers RNAi. This silences essential nematode genes when it feeds on the host, leading to nematode death or impaired survival. The process demonstrates gene-specific silencing mediated by RNA interference, effectively protecting the host. The explanation confirms that the assertion about nematode inability to survive is correctly explained by the RNAi mechanism, highlighting the use of RNAi in controlling parasitic nematodes in transgenic crops.

1. Single Correct Answer MCQ:
Which molecule is directly responsible for nematode gene silencing in a transgenic host?
1. DNA
2. Messenger RNA (mRNA)
3. Interfering RNA (siRNA)
4. Ribosomal RNA (rRNA)
Explanation: Correct answer is interfering RNA (siRNA). Double-stranded RNA derived from sense and antisense transcripts is processed into siRNA. These siRNAs bind complementary nematode mRNA, triggering degradation and silencing gene expression. This mechanism prevents nematode survival when feeding on the host.

2. Single Correct Answer MCQ:
What type of genetic modification is applied in the host to induce nematode resistance?
1. Gene deletion
2. Transgene expression
3. Chromosomal inversion
4. Random mutagenesis
Explanation: Correct answer is transgene expression. A nematode-specific gene is introduced into the host genome, producing interfering RNA. This transgenic approach ensures expression of RNA molecules that induce RNA interference, effectively silencing target nematode genes and providing resistance.

3. Single Correct Answer MCQ:
Which form of RNA is generated from the host to trigger RNAi in nematodes?
1. tRNA
2. Double-stranded RNA
3. Single-stranded mRNA only
4. rRNA
Explanation: Correct answer is double-stranded RNA. Sense and antisense transcripts from the transgene anneal to form dsRNA, which is essential for RNA interference. This dsRNA is processed by Dicer into siRNA, leading to degradation of complementary nematode mRNA and preventing nematode development.

4. Single Correct Answer MCQ:
RNAi mechanism in nematode control is an example of:
1. Protein-mediated defense
2. Host-mediated gene silencing
3. Chemical pesticide application
4. Hormonal regulation
Explanation: Correct answer is host-mediated gene silencing. The transgenic host produces RNA molecules targeting nematode genes. This silences essential nematode genes during feeding. RNA interference is a post-transcriptional gene silencing mechanism leveraged by genetically engineered hosts to confer resistance against parasitic nematodes.

5. Single Correct Answer MCQ:
Which component of RNAi ensures specificity against nematode genes?
1. siRNA sequence complementarity
2. Ribosomal subunits
3. DNA methylation
4. Protein translation factors
Explanation: Correct answer is siRNA sequence complementarity. The siRNA derived from double-stranded RNA is complementary to target nematode mRNA, ensuring gene-specific silencing. Only mRNAs matching the siRNA are degraded, preventing off-target effects while conferring effective nematode resistance.

6. Single Correct Answer MCQ:
Which parasitic organism is controlled using host-mediated RNAi in this context?
1. Fungi
2. Nematode
3. Bacteria
4. Virus
Explanation: Correct answer is nematode. Parasitic nematodes feed on plant roots, causing severe crop losses. By expressing nematode-specific interfering RNA in transgenic hosts, RNAi silences essential nematode genes, preventing survival and proliferation, providing an effective biotechnological control method.

7. Assertion-Reason MCQ:
Assertion (A): Transgenic plants expressing dsRNA can control nematode infections.
Reason (R): dsRNA is processed into siRNA, which degrades complementary nematode mRNA.
1. Both A and R are true and R is the correct explanation of A
2. Both A and R are true but R is not the correct explanation of A
3. A is true, R is false
4. A is false, R is true
Explanation: Correct answer is option 1. The dsRNA expressed by the host is processed into siRNA, which targets nematode mRNA for degradation. This mechanism explains why transgenic plants expressing dsRNA effectively prevent nematode survival.

8. Matching Type MCQ:
Match the component with its function:
A. dsRNA – (i) Triggers RNAi
B. siRNA – (ii) Guides degradation of target mRNA
C. Nematode mRNA – (iii) Target molecule
D. Transgenic host – (iv) Produces dsRNA
1. A–i, B–ii, C–iii, D–iv
2. A–ii, B–i, C–iv, D–iii
3. A–iv, B–iii, C–ii, D–i
4. A–iii, B–iv, C–i, D–ii
Explanation: Correct answer is option 1. dsRNA triggers RNAi, siRNA guides degradation of complementary nematode mRNA, nematode mRNA is the target, and the transgenic host produces dsRNA.

9. Fill in the Blanks MCQ:

Topic: Recombinant DNA Technology; Subtopic: Vectors in rDNA Technology

Keyword Definitions:

• Vector: A DNA molecule used to carry foreign genetic material into a host cell for replication and expression.

• Origin of Replication (ori): A DNA sequence that allows the vector to replicate independently within the host cell.

• Restriction Sites: Specific sequences recognized by restriction enzymes to cut DNA for insertion of foreign genes.

• Marker Genes: Genes that confer selectable traits, such as antibiotic resistance, to identify successful transformations.

• High Copy Number: The number of copies of the vector present in a host cell; higher numbers improve gene expression and plasmid yield.

Lead Question – 2022 (Abroad)
Identify the properties of a good vector used in rDNA technology:
(a) It should have origin of replication supporting a high copy number
(b) It should have preferably more than 2 recognition sites
(c) The restriction sites in vector should be in the antibiotic-resistant genes
(d) It should have suitable marker genes
(e) It should be easy to isolate and purify
Choose the most appropriate answer from the options given below:
1. (a), (c) and (e) only
2. (c), (d) and (e) only
3. (a), (b) and (c) only
4. (a), (c), (d) and (e) only

Explanation:
The correct answer is (a), (c), (d) and (e) only. A good vector should have an origin of replication (ori) to ensure high copy number, making DNA amplification efficient. Restriction sites should avoid marker genes to prevent disruption. Suitable marker genes allow selection of transformed cells. The vector should also be easy to isolate and purify for downstream applications. Having more than two recognition sites is not necessary and can complicate cloning. Therefore, properties (a), (c), (d), and (e) are essential characteristics for a vector in recombinant DNA technology, ensuring efficiency, stability, and ease of use.

1. Which of the following is an essential feature of a cloning vector?
1. High copy number origin of replication
2. Multiple restriction sites in marker gene
3. Very large size (>50 kb)
4. Lack of selectable markers
Explanation: The correct answer is High copy number origin of replication. This ensures that the vector can replicate independently in the host cell, producing multiple copies. Large size makes manipulation difficult, restriction sites should avoid marker genes, and selectable markers are necessary for identifying successful clones. High copy number vectors increase efficiency of cloning and gene expression.

2. Selectable marker genes in vectors are used to:
1. Cut DNA
2. Identify successfully transformed cells
3. Increase plasmid size
4. Inhibit replication
Explanation: Correct answer is Identify successfully transformed cells. Marker genes, such as antibiotic resistance genes, allow only cells that have taken up the vector to grow in selective media. This ensures efficient screening. Marker genes are not used for DNA cutting, increasing plasmid size unnecessarily, or inhibiting replication. They are critical for identifying and isolating recombinant clones.

3. Restriction sites in a vector should:
1. Be present in the marker gene
2. Avoid marker genes
3. Only occur once
4. Always be methylated
Explanation: Correct answer is Avoid marker genes. Inserting foreign DNA into restriction sites located within marker genes can disrupt selectable functions, preventing identification of transformed cells. Properly located restriction sites allow cloning without compromising marker gene expression. While multiple sites are useful, they must not disrupt essential genes. Methylation is not required for all cloning vectors.

4. Why should a vector be easy to isolate and purify?
1. To improve transformation efficiency
2. To ensure accurate restriction digestion
3. For downstream applications like sequencing
4. All of the above
Explanation: Correct answer is All of the above. Ease of isolation and purification ensures sufficient quality and quantity of plasmid DNA for transformation, restriction digestion, and downstream applications such as cloning, sequencing, or expression. Difficult-to-purify vectors reduce efficiency and reproducibility, making this property essential in rDNA technology.

5. A vector with high copy number is preferred because:
1. It is more stable
2. It allows production of many DNA copies
3. It prevents restriction digestion
4. It reduces plasmid size
Explanation: Correct answer is It allows production of many DNA copies. High copy number vectors replicate independently, producing multiple copies per cell, increasing yield of recombinant DNA. This facilitates cloning, gene expression, and analysis. Stability, restriction digestion, and plasmid size are secondary considerations compared to efficient replication and high yield.

6. Which property is NOT necessary for a good vector?
1. Origin of replication
2. Selectable marker genes
3. Multiple restriction sites in marker genes
4. Ease of purification
Explanation: Correct answer is Multiple restriction sites in marker genes. Restriction sites within marker genes can disrupt their function. Essential properties include ori for replication, selectable markers for screening, and ease of purification for downstream processes. Multiple restriction sites should be present outside marker genes to facilitate cloning without affecting selection.

7. Assertion-Reason Type:
Assertion (A): Good vectors must have marker genes.
Reason (R): Marker genes allow selection of transformed cells.
1. Both A and R are true, R is the correct explanation of A
2. Both A and R are true, R is not the correct explanation
3. A is true, R is false
4. A is false, R is true
Explanation: Correct answer is option 1. Marker genes, such as antibiotic resistance genes, enable researchers to identify cells that have taken up the vector. This is an essential property of cloning vectors, ensuring efficient screening and selection. Without markers, transformed cells cannot be distinguished from untransformed ones, making the vector ineffective.

8. Matching Type:
Match the vector property with its purpose:
A. Origin of replication – (i) Copy number control
B. Marker gene – (ii) Selection
C. Ease of purification – (iii) Downstream processing
D. Proper restriction sites – (iv) Insertion of foreign DNA
1. A–i, B–ii, C–iii, D–iv
2. A–ii, B–i, C–iv, D–iii
3. A–iv, B–iii, C–ii, D–i
4. A–i, B–iv, C–ii, D–iii
Explanation: Correct answer is option 1. Origin of replication ensures high copy number, marker genes allow selection, ease of purification facilitates downstream processing, and restriction sites enable insertion of foreign DNA. Proper design of vectors ensures efficient cloning, replication, and identification of recombinant cells in rDNA technology.

9. Fill in the Blanks:
A vector that can replicate independently in the host cell due to its ___________ is ideal for rDNA work.
1. Marker gene
2. Restriction site
3. Origin of replication
4. Promoter
Explanation: Correct answer is Origin of replication. The origin allows the vector to replicate independently, producing multiple copies per cell. This is essential for high-yield cloning, ensuring sufficient recombinant DNA for downstream applications. Marker genes and restriction sites are important but do not control replication.

10. Choose the Correct Statements:
Statement I: A good vector must have selectable marker genes.
Statement II: Restriction sites should be located within marker genes.
1. Statement I correct, Statement II incorrect
2. Statement I incorrect, Statement II correct
3. Both correct
4. Both incorrect
Explanation: Correct answer is Statement I correct, Statement II incorrect. Marker genes allow selection of transformed cells, an essential property. Restriction sites should avoid marker genes to prevent disruption. Proper placement of restriction sites and presence of selectable markers are critical for efficient cloning and vector function in recombinant DNA technology.

Topic: DNA Replication and Amplification; Subtopic: Polymerase Chain Reaction (PCR)

Keyword Definitions:

• DNA helicase: Enzyme that unwinds and separates the double-stranded DNA during replication in cells.

• PCR (Polymerase Chain Reaction): Technique used to amplify a specific DNA segment in vitro using heat and enzymes.

• Taq DNA polymerase: Heat-stable enzyme that synthesizes new DNA strands in PCR.

• Primers: Short DNA sequences that initiate DNA synthesis during PCR.

• Denaturation: The step in PCR where high temperature separates double-stranded DNA into single strands.

• Annealing: Step where primers bind to complementary DNA sequences.

• Extension: Step where new DNA strands are synthesized by Taq polymerase.

• Thermal cycler: Machine that automatically regulates temperature changes during PCR.

Lead Question - 2022 (Abroad)
In a cell, the separation of DNA strands is brought about by the enzyme DNA helicase, whereas in PCR, the separation of DNA strands is due to:
1. High temperature
2. Two sets of Primers
3. Taq DNA polymerase
4. Deoxynucleotides
Explanation: The correct answer is High temperature. In PCR, double-stranded DNA separates during the denaturation step at about 94–98°C, replacing the role of helicase used in cellular replication. Heat breaks the hydrogen bonds between complementary bases. After strand separation, primers bind during annealing, and Taq polymerase extends new DNA strands. This temperature-dependent mechanism enables rapid DNA amplification without enzymes for unwinding. PCR revolutionized molecular biology by allowing DNA replication in vitro for diagnostics, research, and forensic studies. High temperature thus acts as a functional equivalent of helicase during the PCR denaturation process, initiating DNA synthesis efficiently.

1. Single Correct Answer Type:
Which of the following steps of PCR involves the action of Taq DNA polymerase?
1. Denaturation
2. Annealing
3. Extension
4. Cooling
Explanation: The correct answer is Extension. During the extension step (usually at 72°C), Taq DNA polymerase adds nucleotides to the 3' end of primers, synthesizing new DNA strands complementary to the template. This thermostable enzyme, derived from Thermus aquaticus, remains active at high temperatures, making it ideal for PCR. Denaturation separates DNA strands, annealing allows primer binding, and extension completes strand synthesis. Each cycle doubles the DNA quantity, leading to exponential amplification. The heat stability and efficiency of Taq polymerase are key to PCR’s success in molecular diagnostics, cloning, and genetic research applications worldwide.

2. Single Correct Answer Type:
Which temperature step in PCR ensures primer binding to the DNA template?
1. 94°C
2. 72°C
3. 55°C
4. 98°C
Explanation: The correct answer is 55°C. The annealing step occurs around 55°C, where primers bind to complementary DNA sequences on the single-stranded templates. Proper primer annealing is essential for successful amplification because it determines the accuracy and efficiency of the extension phase. If temperature is too high, primers fail to bind; too low, non-specific binding occurs. After annealing, Taq polymerase extends the primers at 72°C. PCR cycles of denaturation, annealing, and extension are repeated multiple times, leading to exponential DNA amplification crucial for diagnostics, genetic testing, and research involving DNA sequencing or cloning.

3. Single Correct Answer Type:
Taq DNA polymerase is preferred in PCR because:
1. It is active at low temperature
2. It is heat-stable
3. It requires helicase
4. It denatures easily
Explanation: The correct answer is It is heat-stable. Taq DNA polymerase, derived from the thermophilic bacterium Thermus aquaticus, remains active even at high temperatures (~72°C). This property allows DNA synthesis during PCR’s repeated heating and cooling cycles without enzyme degradation. Ordinary polymerases would denature during the high-temperature denaturation step. The enzyme’s stability enables automation and efficiency in DNA amplification, critical for forensic analysis, medical diagnostics, and cloning. Its ability to function through numerous cycles without replenishment makes it ideal for rapid, large-scale amplification of specific DNA fragments under controlled thermal cycling conditions.

4. Single Correct Answer Type:
In PCR, the region to be amplified is determined by:
1. Primers
2. Template DNA
3. Taq DNA polymerase
4. Nucleotides
Explanation: The correct answer is Primers. Short oligonucleotide sequences called primers define the start and end points of the DNA region to be amplified. They bind to specific complementary sequences on the template DNA during the annealing step. Once bound, Taq polymerase extends the primers to synthesize new DNA strands. The specificity of primers ensures only the target DNA segment is replicated. Incorrect primer design may lead to non-specific amplification. Hence, primers play a vital role in ensuring accuracy, efficiency, and reproducibility of PCR-based experiments, including genetic testing, diagnostics, and research applications in molecular biology laboratories.

5. Single Correct Answer Type:
The enzyme DNA helicase is required during:
1. Transcription
2. PCR
3. In vivo DNA replication
4. RNA processing
Explanation: The correct answer is In vivo DNA replication. DNA helicase is an enzyme that unwinds and separates the double-stranded DNA helix into single strands during replication in living cells. This process creates replication forks where DNA polymerase can synthesize new strands. In PCR, high temperature replaces helicase’s function by denaturing DNA. Helicase activity ensures accurate and efficient replication by enabling template accessibility for polymerase enzymes. Its role is crucial in maintaining genomic integrity and facilitating repair mechanisms. In contrast, PCR relies on controlled heat cycles instead of enzymatic unwinding for DNA strand separation and amplification.

6. Single Correct Answer Type:
Which of the following is NOT required for PCR?
1. Template DNA
2. DNA helicase
3. Primers
4. Taq polymerase
Explanation: The correct answer is DNA helicase. Unlike cellular replication, PCR does not require helicase for unwinding DNA strands. Instead, high temperature denatures DNA during the initial step, allowing primer binding and synthesis by Taq polymerase. Template DNA provides the sequence to be amplified, primers determine specificity, and Taq polymerase catalyzes new strand formation. The absence of helicase simplifies the process, enabling in vitro DNA amplification using thermal cycling. This automation allows replication of specific genes or DNA fragments efficiently for applications in diagnostics, genetic engineering, forensics, and evolutionary studies, eliminating the need for complex enzymatic systems.

7. Assertion-Reason Type:
Assertion (A): PCR can amplify DNA without helicase.
Reason (R): High temperature during denaturation separates the DNA strands.
1. Both A and R are correct, and R explains A
2. Both A and R are correct, but R does not explain A
3. A is correct, R is incorrect
4. A is incorrect, R is correct
Explanation: The correct answer is Both A and R are correct, and R explains A. In PCR, high temperature (94–98°C) replaces helicase by denaturing DNA, breaking hydrogen bonds between strands. This thermal denaturation makes DNA single-stranded for primer annealing. Unlike in vivo replication requiring helicase, PCR uses physical heat for strand separation. The thermal cycling method enables repeated denaturation and synthesis cycles for DNA amplification. The principle behind PCR’s success lies in substituting biological mechanisms with heat, simplifying the process while maintaining efficiency, making it an essential tool in modern molecular biology, diagnostics, and forensic applications.

8. Matching Type:
Match the PCR step with its function:
A. Denaturation → (i) Primer binding
B. Annealing → (ii) Strand separation
C. Extension → (iii) DNA synthesis
Options:
1. A-ii, B-i, C-iii
2. A-i, B-ii, C-iii
3. A-iii, B-i, C-ii
4. A-ii, B-iii, C-i
Explanation: The correct answer is 1. A-ii, B-i, C-iii. Denaturation separates double-stranded DNA (A-ii), annealing allows primer binding (B-i), and extension synthesizes new DNA strands (C-iii). Each PCR cycle repeats these three steps, leading to exponential DNA amplification. Denaturation typically occurs at 94–98°C, annealing at 50–60°C, and extension at 72°C. These temperature-dependent reactions ensure specificity, precision, and high efficiency. The cyclical nature of PCR allows billions of copies of DNA fragments to be produced within hours, making it indispensable for genetic testing, cloning, disease detection, and forensic analysis in molecular biology laboratories globally.

9. Fill in the Blanks Type:
In PCR, the enzyme used for synthesis of DNA is __________.
1. DNA helicase
2. DNA ligase
3. Taq polymerase
4. RNA polymerase
Explanation: The correct answer is Taq polymerase. Taq polymerase, derived from the thermophilic bacterium Thermus aquaticus, synthesizes DNA during the extension phase of PCR. Its heat stability enables it to function at high temperatures, avoiding degradation during denaturation cycles. This enzyme extends primers by adding nucleotides complementary to the DNA template. The use of Taq polymerase made PCR efficient and automated, revolutionizing molecular biology by allowing DNA amplification for various applications like disease diagnostics, cloning, sequencing, and forensic analysis. It remains one of the most essential enzymes in biotechnology and genetic research worldwide.

10. Choose the Correct Statements (Statement I & II):
Statement I: DNA helicase is required for strand separation during PCR.
Statement II: High temperature acts as a substitute for helicase in PCR.
1. Both statements are correct
2. Both statements are incorrect
3. Statement I is incorrect, but Statement II is correct
4. Statement I is correct, but Statement II is incorrect
Explanation: The correct answer is Statement I is incorrect, but Statement II is correct. In PCR, high temperature (~95°C) denatures DNA strands by breaking hydrogen bonds, eliminating the need for helicase. Helicase functions only in biological systems for DNA unwinding during replication. PCR’s thermal process mimics this separation, allowing primers to anneal and Taq polymerase to synthesize new strands. This simplicity and reliability make PCR a revolutionary method for in vitro DNA amplification. Understanding the distinction between enzymatic and thermal strand separation clarifies why PCR efficiently mimics natural DNA replication in controlled laboratory environments.

Topic: Recombinant DNA Technology; Subtopic: Plasmids and Restriction Enzymes

Keyword Definitions:

• Plasmid: A small, circular, double-stranded DNA molecule found in bacteria, separate from chromosomal DNA.

• Recombinant DNA: DNA molecule formed by combining genetic material from two different sources.

• Restriction Enzymes: Molecular scissors that cut DNA at specific sequences.

• Host Organism: The bacterial species into which recombinant DNA is introduced for replication or expression.

• Antibiotic Resistance Gene: A gene that provides bacteria resistance to antibiotics, used as a marker in cloning.

Lead Question - 2022 (Abroad)
The construction of the first recombinant DNA emerged from the possibility of linking a gene encoding antibiotic resistance with a native plasmid of which of the following organism?
1. Escherichia coli
2. Bacillus thuringiensis
3. Salmonella typhimurium
4. Agrobacterium tumefaciens
Explanation: The first recombinant DNA was created by combining antibiotic resistance gene with the plasmid of Salmonella typhimurium using restriction enzyme EcoRI. This discovery by Stanley Cohen and Herbert Boyer in 1973 marked the foundation of genetic engineering. The plasmid used was pSC101, and this pioneering experiment led to the birth of recombinant DNA technology.

1. Single Correct Answer Type:
Which of the following enzymes is essential for joining foreign DNA with plasmid DNA?
1. RNA polymerase
2. DNA ligase
3. DNA helicase
4. Restriction endonuclease
Explanation: The enzyme DNA ligase acts as a molecular glue that seals the phosphodiester backbone between foreign DNA and plasmid DNA. This enzyme ensures stable recombinant DNA formation, allowing replication inside the host cell. It is derived from bacteriophages and is vital in DNA cloning and molecular biotechnology applications.

2. Single Correct Answer Type:
Which restriction enzyme was first used to produce recombinant DNA?
1. HindIII
2. BamHI
3. EcoRI
4. PvuII
Explanation: The restriction enzyme EcoRI was used by Cohen and Boyer in 1973 to cut DNA at specific recognition sequences, producing sticky ends. These sticky ends allowed insertion of foreign DNA fragments into plasmids, leading to successful recombinant DNA construction. EcoRI recognizes the sequence GAATTC and produces overhanging ends for ligation.

3. Single Correct Answer Type:
In recombinant DNA technology, the plasmid used as vector should have
1. No selectable marker
2. Only a single origin of replication
3. Multiple replication origins
4. No restriction site
Explanation: A good cloning vector should have a single origin of replication (ori), which ensures controlled and efficient replication inside the host cell. It should also contain selectable markers, such as antibiotic resistance genes, and multiple cloning sites for insertion of foreign DNA without affecting vector stability.

4. Single Correct Answer Type:
Which of the following is used as a selectable marker in plasmid pBR322?
1. Ampicillin and tetracycline resistance genes
2. Kanamycin and chloramphenicol resistance genes
3. Streptomycin and neomycin resistance genes
4. Rifampicin and erythromycin resistance genes
Explanation: The plasmid pBR322 contains two antibiotic resistance genes – one for ampicillin and another for tetracycline. These serve as selectable markers to identify recombinant colonies after transformation. The plasmid was developed by Bolivar and Rodriguez and is one of the most commonly used vectors in biotechnology laboratories worldwide.

5. Single Correct Answer Type:
Which of the following is not a property of an ideal cloning vector?
1. Presence of origin of replication
2. Presence of multiple cloning site
3. Large size of plasmid
4. Presence of selectable marker
Explanation: An ideal cloning vector must be small in size for easy manipulation. Hence, large plasmid size is not a desirable feature. Small vectors replicate faster, are more stable, and easier to isolate. Other properties like ori, selectable markers, and restriction sites are crucial for successful recombinant DNA cloning.

6. Single Correct Answer Type:
Which organism serves as a natural genetic engineer due to its Ti plasmid?
1. Agrobacterium tumefaciens
2. Escherichia coli
3. Bacillus subtilis
4. Pseudomonas aeruginosa
Explanation: Agrobacterium tumefaciens is known as a natural genetic engineer because it transfers a segment of its Ti (tumor-inducing) plasmid into plant genomes, causing crown gall disease. Scientists modify this Ti plasmid to transfer desirable genes, making it a powerful tool in genetic transformation of plants for crop improvement.

7. Assertion-Reason Type:
Assertion (A): The enzyme EcoRI cuts DNA producing sticky ends.
Reason (R): Sticky ends facilitate the joining of DNA fragments by complementary base pairing.
1. Both A and R are true, and R is the correct explanation of A.
2. Both A and R are true, but R is not the correct explanation of A.
3. A is true, but R is false.
4. A is false, but R is true.
Explanation: Both Assertion and Reason are true, and the Reason correctly explains the Assertion. EcoRI produces sticky ends that can base-pair with complementary sequences on other DNA molecules, enabling effective ligation by DNA ligase to form stable recombinant DNA molecules during cloning experiments.

8. Matching Type:
Match the following:
A. EcoRI → (i) Sticky ends
B. HindII → (ii) Blunt ends
C. DNA ligase → (iii) Joining DNA fragments
D. Plasmid → (iv) Vector
1. A-(i), B-(ii), C-(iii), D-(iv)
2. A-(ii), B-(i), C-(iii), D-(iv)
3. A-(i), B-(ii), C-(iv), D-(iii)
4. A-(iii), B-(i), C-(ii), D-(iv)
Explanation: The correct matching is A-(i), B-(ii), C-(iii), D-(iv). EcoRI produces sticky ends, HindII generates blunt ends, DNA ligase joins DNA fragments, and plasmids act as vectors. These components collectively play a key role in forming recombinant DNA molecules in molecular cloning experiments.

9. Fill in the Blanks Type:
The enzyme used to cut DNA molecules at specific sites is called ________.
1. Polymerase
2. Ligase
3. Restriction endonuclease
4. Transcriptase
Explanation: The correct answer is Restriction endonuclease. These enzymes recognize specific palindromic DNA sequences and cleave them to produce fragments with sticky or blunt ends. They are essential for generating DNA fragments that can be recombined with vectors in genetic engineering for gene cloning and molecular diagnostics.

10. Choose the Correct Statements Type:
Choose the correct statements regarding recombinant DNA technology.
1. Restriction enzymes cut DNA at random sites.
2. DNA ligase joins DNA fragments.
3. Plasmids act as carriers of foreign DNA.
4. Both 2 and 3 are correct.
Explanation: The correct choice is Both 2 and 3 are correct. DNA ligase joins fragments by forming phosphodiester bonds, and plasmids serve as cloning vectors carrying foreign genes into host cells. Together, they are essential tools in genetic engineering for developing recombinant organisms and biotechnological products.

Subtopic: rDNA Technology and Selection

• rDNA Technology: Recombinant DNA technology involves combining DNA from different sources to create genetically modified organisms.

• Non-recombinant bacteria: Bacteria that do not take up the recombinant plasmid and retain original traits.

• Recombinant Plasmid: Circular DNA molecule engineered to carry a foreign gene for transformation into host bacteria.

• Transformation: Process by which bacteria take up foreign DNA, including recombinant plasmids, from the environment.

• Foreign Gene: Gene introduced into an organism from a different species to confer new traits.

• Ampicillin: Antibiotic used as a selectable marker in plasmids to identify transformed bacteria.

• Tetracycline: Antibiotic used in rDNA experiments for selection of recombinant or non-recombinant bacteria.

• Selectable Marker: Gene in a plasmid conferring resistance to an antibiotic, enabling identification of transformed cells.

• Host Bacteria: Bacterial strain used to propagate recombinant plasmids.

• Cloning Vector: DNA molecule (plasmid) used to carry foreign genes into host cells.

• Gene of Interest: Specific foreign gene intended to be expressed in host bacteria using rDNA technology.

Lead Question - 2022 (Abroad)

Given below are two statements: One is labelled as Assertion (A) and the other is labelled as Reason (R).

Assertion (A): In rDNA technology non-recombinants transformed bacteria grow on the medium containing ampicillin as well as medium containing tetracycline.

Reason (R): Recombinant plasmids contain the foreign gene of interest.

In the light of the above statements, choose the correct answer from the options given below:

1. (A) is correct but (R) is not correct

2. (A) is not correct but (R) is correct

3. Both (A) and (R) are correct and (R) is the correct explanation of (A)

4. Both (A) and (R) are correct but (R) is not the correct explanation of (A)

Explanation: In rDNA technology, non-recombinant bacteria carry plasmids without foreign gene insertion and can grow on both antibiotics if the plasmid has dual resistance markers. Recombinant plasmids carry the gene of interest but may lose one marker. Hence, assertion is correct but reason does not explain it. Answer: (A) is correct but (R) is not correct. Answer: 1

Q1: Which enzyme is used to cut DNA at specific sites to create recombinant plasmids?

1. DNA polymerase

2. RNA polymerase

3. Restriction endonuclease

4. Ligase

Explanation: Restriction endonucleases cut DNA at specific recognition sequences, generating sticky or blunt ends that allow insertion of foreign genes into plasmids. DNA polymerase synthesizes DNA, RNA polymerase synthesizes RNA, and ligase joins DNA fragments. Answer: Restriction endonuclease. Answer: 3

Q2: Which is a common selectable marker used in recombinant plasmids?

1. Green fluorescent protein

2. Ampicillin resistance gene

3. Lac operon

4. Origin of replication

Explanation: Ampicillin resistance gene allows identification of transformed bacteria by surviving on ampicillin-containing medium. GFP is a reporter, Lac operon regulates transcription, and origin of replication is essential for plasmid replication. Answer: Ampicillin resistance gene. Answer: 2

Q3: What is the role of ligase in rDNA technology?

1. Cut DNA at specific sites

2. Join DNA fragments

3. Replicate plasmid

4. Insert plasmid into bacteria

Explanation: DNA ligase catalyzes the formation of phosphodiester bonds between DNA fragments, joining foreign DNA and plasmid to create recombinant DNA. Cutting is done by restriction endonucleases, replication occurs via host machinery, and plasmid insertion is by transformation. Answer: Join DNA fragments. Answer: 2

Q4: Which method is used to introduce recombinant plasmid into bacteria?

1. Transcription

2. Transformation

3. Translation

4. Transduction

Explanation: Transformation is the process of introducing recombinant DNA into host bacteria. Transcription and translation are gene expression processes, while transduction is viral-mediated gene transfer. Answer: Transformation. Answer: 2

Q5: What type of plasmid is commonly used in cloning foreign genes?

1. Conjugative plasmid

2. Fertility plasmid

3. Cloning vector

4. Resistance plasmid

Explanation: Cloning vectors are plasmids engineered to carry foreign genes for replication and expression in host cells. Conjugative plasmids mediate transfer, fertility plasmids enable mating, and resistance plasmids carry antibiotic resistance. Answer: Cloning vector. Answer: 3

Q6: Which bacterial strain is preferred as host for rDNA experiments?

1. Escherichia coli

2. Bacillus subtilis

3. Staphylococcus aureus

4. Pseudomonas putida

Explanation: E. coli is widely used as host due to well-characterized genetics, rapid growth, and ability to maintain plasmids. Other bacteria are less suitable for general cloning. Answer: Escherichia coli. Answer: 1

Q7: Assertion (A): Antibiotic resistance genes are essential in recombinant plasmids.
Reason (R): They help in identifying and selecting transformed bacteria.

1. A is correct but R is not correct

2. A is not correct but R is correct

3. Both A and R are correct and R explains A

4. Both A and R are correct but R does not explain A

Explanation: Antibiotic resistance genes act as selectable markers, allowing only bacteria that contain the recombinant plasmid to survive on antibiotic media. This directly explains their importance in recombinant plasmids. Answer: Both A and R are correct and R explains A. Answer: 3

Q8: Match plasmid types with their roles:

1. Cloning vector    A. Carry foreign genes
2. Reporter plasmid    B. Detect expression
3. Resistance plasmid    C. Select transformed cells
4. Conjugative plasmid    D. Transfer DNA between bacteria

1. 1-A, 2-B, 3-C, 4-D

2. 1-B, 2-A, 3-D, 4-C

3. 1-C, 2-D, 3-A, 4-B

4. 1-D, 2-C, 3-B, 4-A

Explanation: Cloning vectors carry foreign genes, reporter plasmids detect gene expression, resistance plasmids help select transformed cells, and conjugative plasmids transfer DNA between bacteria. Correct match: 1-A, 2-B, 3-C, 4-D. Answer: 1

Q9: A gene of interest is inserted into a plasmid to create a ______ plasmid.

1. Recombinant

2. Resistance

3. Conjugative

4. Reporter

Explanation: Inserting a foreign gene into a plasmid produces a recombinant plasmid that can replicate and express the inserted gene in host bacteria. Resistance, conjugative, and reporter plasmids have different primary functions. Answer: Recombinant. Answer: 1

Q10: Select correct statements about rDNA technology:

1. Uses restriction enzymes to cut DNA

2. Ligase joins DNA fragments

3. Selectable markers allow identification of transformants

4. Transformation introduces plasmids into host bacteria

Explanation: rDNA technology involves cutting DNA with restriction enzymes, joining fragments with ligase, using selectable markers to identify transformed bacteria, and introducing plasmids via transformation. All listed statements accurately describe fundamental steps in recombinant DNA experiments. Answer: 1,2,3,4</

Subtopic: Restriction Enzymes

• Restriction Enzyme: An enzyme that cuts DNA at specific nucleotide sequences, also called restriction endonuclease.

• Endonuclease: Enzymes that cleave the phosphodiester bond within a DNA or RNA molecule.

• DNA Cleavage: The process of breaking DNA strands at specific sites.

• Specific Sites: Particular nucleotide sequences recognized by restriction enzymes for cutting DNA.

• Double-Stranded DNA: DNA composed of two complementary strands forming a helix.

• Palindromic Sequence: DNA sequence that reads the same in 5' to 3' direction on both strands, recognized by restriction enzymes.

• Sticky Ends: Overhanging single-stranded ends produced by restriction enzyme cleavage.

• Blunt Ends: DNA ends without overhangs after restriction enzyme cutting.

• Molecular Cloning: Technique using restriction enzymes to insert DNA fragments into vectors.

• Recombinant DNA: DNA molecule formed by combining DNA from different sources.

• Genetic Engineering: Use of restriction enzymes and other tools to manipulate DNA for research or biotechnology applications.

Lead Question - 2022 (Abroad)

Given below are two statements: One is labelled as Assertion (A) and the other is labelled as Reason (R)
Assertion (A): Restriction enzyme is a type of endonuclease
Reason (R) : Restriction enzyme cuts the two strands of DNA at specific positions within the DNA

1. (A) is correct but (R) is not correct

2. (A) is not correct but (R) is correct

3. Both (A) and (R) are correct and (R) is the correct explanation of (A)

4. Both (A) and (R) are correct but (R) is not the correct explanation of (A)

Explanation: Restriction enzymes are indeed endonucleases because they cleave phosphodiester bonds within DNA strands. They recognize specific palindromic sequences and cut both strands of DNA at precise positions, creating sticky or blunt ends. Therefore, both Assertion (A) and Reason (R) are correct, and R explains A. Answer: 3

Q1: Which type of ends can be generated by restriction enzyme cleavage?

1. Sticky ends only

2. Blunt ends only

3. Both sticky and blunt ends

4. Neither sticky nor blunt ends

Explanation: Restriction enzymes can cut DNA in a manner that produces sticky ends with overhangs or blunt ends with no overhangs. The type of end depends on the enzyme used. This feature is essential for recombinant DNA techniques. Answer: Both sticky and blunt ends. Answer: 3

Q2: Restriction enzymes recognize which type of DNA sequence?

1. Random sequence

2. Promoter sequence

3. Palindromic sequence

4. Terminator sequence

Explanation: Restriction enzymes specifically recognize palindromic sequences in DNA, which read the same forwards and backwards on complementary strands. This specificity allows precise cutting for molecular cloning and recombinant DNA experiments. Answer: Palindromic sequence. Answer: 3

Q3: Restriction enzymes are classified as:

1. Exonucleases

2. Endonucleases

3. Ligases

4. Polymerases

Explanation: Restriction enzymes are endonucleases because they cleave the phosphodiester bonds within DNA strands rather than at terminal ends. Exonucleases remove nucleotides from ends, while ligases join DNA fragments. Answer: Endonucleases. Answer: 2

Q4: Which application uses restriction enzymes?

1. Molecular cloning

2. Polymerase chain reaction

3. Gel electrophoresis

4. DNA sequencing

Explanation: Restriction enzymes are crucial in molecular cloning as they cut DNA at specific sites to allow insertion into vectors, producing recombinant DNA molecules. Other techniques may use DNA but do not require site-specific cleavage. Answer: Molecular cloning. Answer: 1

Q5: What is the main function of restriction enzymes in genetic engineering?

1. Amplify DNA

2. Cut DNA at specific sites

3. Join DNA fragments

4. Transcribe DNA to RNA

Explanation: Restriction enzymes cut DNA at specific recognition sites, producing fragments suitable for ligation and cloning, which is fundamental in genetic engineering and recombinant DNA technology. Amplification is performed by polymerase, not restriction enzymes. Answer: Cut DNA at specific sites. Answer: 2

Q6: Restriction enzymes are essential tools in:

1. Protein folding studies

2. Genetic manipulation

3. Cell membrane synthesis

4. Photosynthesis research

Explanation: Restriction enzymes are used to cut and manipulate DNA sequences, making them indispensable tools for genetic engineering, recombinant DNA technology, and molecular biology experiments. Answer: Genetic manipulation. Answer: 2

Q7: Assertion (A): EcoRI is a type of restriction enzyme.
Reason (R): EcoRI produces sticky ends by cutting DNA at specific sites.

1. A is correct but R is not correct

2. A is not correct but R is correct

3. Both A and R are correct and R explains A

4. Both A and R are correct but R does not explain A

Explanation: EcoRI is a well-known restriction enzyme that recognizes the sequence GAATTC and cuts DNA, producing sticky ends. Both A and R are correct, and R correctly explains A. Answer: Both A and R are correct and R explains A. Answer: 3

Q8: Match restriction enzyme type with its recognition pattern:

1. EcoRI    A. GAATTC
2. HindIII    B. AAGCTT
3. BamHI    C. GGATCC
4. SmaI    D. CCCGGG

1. 1-A, 2-B, 3-C, 4-D

2. 1-B, 2-A, 3-D, 4-C

3. 1-C, 2-D, 3-A, 4-B

4. 1-D, 2-C, 3-B, 4-A

Explanation: EcoRI recognizes GAATTC, HindIII recognizes AAGCTT, BamHI recognizes GGATCC, and SmaI recognizes CCCGGG sequences in DNA. These recognition sites allow specific cleavage for molecular cloning. Correct match: 1-A, 2-B, 3-C, 4-D. Answer: 1

Q9: Restriction enzymes are used to produce _______ for recombinant DNA experiments.

1. RNA fragments

2. DNA fragments

3. Proteins

4. Polysaccharides

Explanation: Restriction enzymes cut DNA at specific sequences, generating DNA fragments. These fragments can be ligated into vectors for recombinant DNA technology and genetic engineering applications. Answer: DNA fragments. Answer: 2

Q10: Select the correct statements about restriction enzymes:

1. They are endonucleases

2. They recognize specific DNA sequences

3. They can produce sticky or blunt ends

4. They are essential for recombinant DNA technology

Explanation: All four statements are correct. Restriction enzymes are endonucleases that cut DNA at specific sequences, produce sticky or blunt ends, and are indispensable tools in recombinant DNA technology and genetic engineering. Answer: 1, 2, 3, 4

ubtopic: Recombinant Protein Production

• Genetically engineered insulin: Insulin produced using recombinant DNA technology for therapeutic use in humans.

• Escherichia coli: A commonly used bacterial host in biotechnology for recombinant protein production.

• Pseudomonas putida: Soil bacterium used in bioremediation, occasionally for genetic studies.

• Bacillus thuringiensis: Bacterium producing insecticidal toxins, used in pest control.

• Rhizobium meliloti: Nitrogen-fixing bacterium associated with legumes.

• Recombinant DNA technology: Technique to combine DNA from different sources to produce desired proteins.

• Host organism: Organism used to produce recombinant proteins.

• Protein expression: Production of protein from introduced gene in a host organism.

• Insulin: Hormone regulating blood glucose, used therapeutically in diabetes.

• Biotechnology: Application of biological systems for industrial, medical, or agricultural purposes.

• Therapeutic proteins: Proteins produced for medical treatment of diseases.

Lead Question - 2022 (Abroad)

Genetically engineered insulin for human is produced from:

1. Escherichia coli

2. Pseudomonas putida

3. Bacillus thuringiensis

4. Rhizobium meliloti

Explanation: Human insulin is produced using recombinant DNA technology in Escherichia coli, which is genetically modified to express insulin gene. This method provides large-scale, safe, and cost-effective insulin. Other bacteria like Pseudomonas, Bacillus, and Rhizobium are not typically used for human insulin production. Answer: Escherichia coli. Answer: 1

Q1: Which of the following is a common host for recombinant protein production?

1. Escherichia coli

2. Bacillus subtilis

3. Yeast

4. All of the above

Explanation: E. coli, Bacillus subtilis, and yeast are commonly used as hosts in recombinant protein production due to their fast growth, easy genetic manipulation, and ability to express proteins efficiently. This versatility allows for therapeutic protein production. Answer: All of the above. Answer: 4

Q2: The insulin gene is inserted into E. coli using:

1. Plasmid vector

2. Viral vector

3. Chromosomal recombination

4. Transposon insertion

Explanation: A plasmid vector carries the human insulin gene into E. coli, enabling the bacterium to produce insulin protein. Plasmids replicate independently, ensuring stable gene expression. Viral vectors and transposons are alternative methods not commonly used for insulin production. Answer: Plasmid vector. Answer: 1

Q3: Which is the first step in recombinant insulin production?

1. Protein purification

2. Gene cloning

3. Bacterial culture growth

4. Folding of insulin

Explanation: Gene cloning is the first step, where the human insulin gene is isolated and inserted into a plasmid vector for bacterial expression. Subsequent steps involve bacterial culture, protein expression, purification, and folding. Answer: Gene cloning. Answer: 2

Q4: What is the advantage of recombinant insulin over animal-derived insulin?

1. Cheaper production

2. Reduced immune reaction

3. Large-scale production

4. All of the above

Explanation: Recombinant insulin is produced cheaply in large amounts, is identical to human insulin, and reduces immune reactions compared to animal insulin. It is safer and more consistent for diabetes treatment. Answer: All of the above. Answer: 4

Q5: In genetic engineering, what ensures insulin gene expression in E. coli?

1. Promoter sequence

2. Origin of replication

3. Selectable marker

4. Restriction enzyme

Explanation: Promoter sequences in the plasmid vector drive transcription of the insulin gene in E. coli. Origin of replication ensures plasmid replication, selectable markers allow selection, and restriction enzymes aid cloning. Expression requires the promoter. Answer: Promoter sequence. Answer: 1

Q6: Which protein modification is sometimes required for recombinant insulin?

1. Glycosylation

2. Disulfide bond formation

3. Phosphorylation

4. Methylation

Explanation: Human insulin has disulfide bonds critical for its function. Recombinant insulin requires correct folding and bond formation. Glycosylation, phosphorylation, or methylation are not needed for insulin. Proper disulfide bond formation ensures biological activity. Answer: Disulfide bond formation. Answer: 2

Q7: Assertion (A): Escherichia coli is preferred for recombinant insulin production.
Reason (R): It grows fast, is easy to manipulate, and produces proteins in large amounts.

1. (A) is correct but R is not correct

2. (A) is not correct but R is correct

3. Both A and R are correct and R explains A

4. Both A and R are correct but R does not explain A

Explanation: E. coli is ideal for recombinant insulin due to rapid growth, easy genetic manipulation, and high protein yield. Both assertion and reason are correct, and reason directly explains the assertion. Answer: Both A and R are correct and R explains A. Answer: 3

Q8: Match the component with its role in recombinant insulin production:

A. Plasmid vector    1. Carry gene
B. Promoter    2. Drive transcription
C. E. coli    3. Host for expression
D. Disulfide bonds    4. Functional protein folding

1. A-1, B-2, C-3, D-4

2. A-2, B-1, C-3, D-4

3. A-1, B-3, C-2, D-4

4. A-4, B-3, C-2, D-1

Explanation: Correct matching: Plasmid vector carries gene (A-1), promoter drives transcription (B-2), E. coli is the host (C-3), disulfide bonds ensure correct folding (D-4). This clarifies recombinant insulin production process. Answer: A-1, B-2, C-3, D-4. Answer: 1

Q9: The process of producing insulin using recombinant DNA in bacteria is called ______.

1. Transgenesis

2. Recombinant protein production

3. Fermentation

4. Cloning

Explanation: Recombinant protein production involves expressing a gene in a host organism to produce the protein. Insulin is produced this way in E. coli. Transgenesis is gene insertion into organisms, fermentation is culture growth, cloning is gene replication. Answer: Recombinant protein production. Answer: 2

Q10: Which statements about recombinant insulin are correct?

1. Produced in E. coli

2. Identical to human insulin

3. Safer than animal insulin

4. Requires glycosylation for activity

Explanation: Recombinant insulin is produced in E. coli, is structurally identical to human insulin, and safer than animal-derived insulin. It does not require glycosylation to be active. Correct statements are 1, 2, 3. Answer: 1, 2, 3

Topic: Genetic Engineering
Subtopic: Cloning Vectors and Recombinant DNA Technology

Keyword Definitions:
- Cloning vector: DNA molecule used to carry foreign DNA into a host cell.
- Marker gene: Gene used to identify cells containing the vector.
- Restriction enzyme site: Specific DNA sequence recognized and cut by an enzyme.
- Origin of replication (ori): DNA sequence allowing replication of vector in host.
- Recombinant DNA: DNA formed by combining sequences from different sources.
- Plasmid: Circular DNA molecule used as a cloning vector.
- Selectable marker: Gene allowing selection of successfully transformed cells.
- Multiple cloning site: DNA region containing several restriction sites.
- Host cell: Cell that receives and replicates the vector.
- Gene cloning: Production of identical copies of a DNA fragment.
- Vector features: Properties such as size, replication origin, and markers ensuring efficient cloning.

Lead Question - 2022:
Which of the following is not a desirable feature of a cloning vector?
(1) Presence of a marker gene
(2) Presence of single restriction enzyme site
(3) Presence of two or more recognition sites
(4) Presence of origin of replication

Explanation: The correct answer is (2). A cloning vector should ideally have multiple recognition sites, not a single site, allowing insertion of foreign DNA. Presence of a marker gene, multiple restriction sites, and origin of replication are desirable features ensuring selection, replication, and versatility in genetic engineering.

1. Single Correct Answer:
Which feature allows a vector to replicate independently in a host?
(a) Marker gene
(b) Origin of replication
(c) Restriction site
(d) Antibiotic resistance
Explanation: The origin of replication (ori) enables independent replication of the vector within the host. Marker genes and antibiotic resistance allow selection but do not support replication. Restriction sites allow DNA insertion but are not responsible for vector replication.

2. Single Correct Answer:
Which type of vector is commonly used for gene cloning in bacteria?
(a) Plasmid
(b) Virus
(c) Cosmid
(d) All of the above
Explanation: Plasmids are the most commonly used bacterial cloning vectors. They are small, circular DNA molecules with origin of replication, marker genes, and multiple cloning sites, allowing easy insertion and replication of foreign DNA. Viruses and cosmids are also used but less frequently.

3. Single Correct Answer:
A selectable marker gene in a vector is used for:
(a) Replication
(b) DNA cutting
(c) Identifying transformed cells
(d) Enhancing transcription
Explanation: Selectable marker genes allow identification of cells that have successfully taken up the vector. These markers, often conferring antibiotic resistance, help distinguish transformed cells from non-transformed ones. They do not directly aid replication or transcription but enable efficient selection in cloning experiments.

4. Single Correct Answer:
Multiple cloning sites in a vector provide:
(a) Antibiotic resistance
(b) Multiple restriction sites for DNA insertion
(c) Replication origin
(d) Marker gene
Explanation: Multiple cloning sites (MCS) contain several restriction enzyme recognition sites, allowing insertion of DNA fragments in various ways. Marker genes, origin of replication, and antibiotic resistance are distinct features. MCS ensures versatility in cloning and flexibility for inserting genes into the vector.

5. Single Correct Answer:
Which of the following is not a vector used in eukaryotic gene cloning?
(a) Plasmid
(b) Bacteriophage
(c) Yeast artificial chromosome
(d) Ti plasmid
Explanation: Bacteriophages are primarily bacterial vectors, not used in eukaryotic gene cloning. Plasmids, yeast artificial chromosomes (YACs), and Ti plasmids are effective in eukaryotic systems. Selecting an appropriate vector ensures efficient replication, DNA insertion, and gene expression in the target host.

6. Single Correct Answer:
Which feature of a vector allows easy identification of recombinant clones?
(a) Origin of replication
(b) Marker gene
(c) Restriction site
(d) Polylinker
Explanation: A marker gene enables identification of recombinant clones. For example, antibiotic resistance genes or reporter genes like lacZ allow selection or visual detection. Origin of replication ensures replication, restriction sites allow insertion, and polylinker provides multiple insertion options but not direct identification.

7. Assertion-Reason MCQ:
Assertion (A): A desirable vector should have multiple recognition sites.
Reason (R): Multiple sites allow insertion of foreign DNA at different positions.
(a) Both A and R are true, R explains A
(b) Both A and R are true, R does not explain A
(c) A is true, R is false
(d) A is false, R is true
Explanation: Option (a) is correct. A vector with multiple restriction sites (MCS) allows insertion of DNA fragments at different positions without disrupting essential vector functions. Both the assertion and reason are correct, and R directly explains why multiple recognition sites are desirable.

8. Matching Type MCQ:
Match vector features with their functions:
List - I List - II
(a) Marker gene (i) Selection of transformed cells
(b) Origin of replication (ii) Independent replication
(c) Multiple cloning site (iii) Multiple DNA insertion points
(d) Restriction enzyme site (iv) DNA cleavage
Options:
(1) a-i, b-ii, c-iii, d-iv
(2) a-ii, b-i, c-iv, d-iii
(3) a-iii, b-iv, c-i, d-ii
(4) a-iv, b-iii, c-ii, d-i
Explanation: Option (1) is correct. Marker genes allow selection, origin of replication enables replication, multiple cloning site provides insertion flexibility, and restriction enzyme site allows DNA cleavage. Proper mapping ensures efficient vector design and gene cloning.

9. Fill in the Blanks:
A cloning vector must contain an _______ to replicate inside the host.
(a) Marker gene
(b) Multiple cloning site
(c) Origin of replication
(d) Restriction enzyme site
Explanation: A cloning vector must contain an origin of replication (ori) to replicate independently in the host cell. Marker genes, restriction sites, and MCS support selection and DNA insertion but cannot drive vector replication on their own.

10. Choose the Correct Statements:
Identify correct statements:
1. A vector must have a selectable marker.
2. Single restriction site is sufficient for all cloning.
3. Multiple cloning sites increase flexibility.
4. Origin of replication is required for vector replication.
Options:
(a) 1, 2, 3
(b) 1, 3, 4
(c) 2, 3, 4
(d) 1, 2, 4
Explanation: Option (b) is correct. Vectors should have a marker for selection, multiple cloning sites for insertion flexibility, and an origin for replication. A single restriction site is not sufficient for versatile cloning, making statement 2 incorrect.

Subtopic: Genetically Engineered Insulin

• Insulin: A peptide hormone produced by pancreatic beta cells that regulates blood glucose levels.

• Pro-hormone Insulin: Immature form of insulin containing extra C-peptide chain.

• C-peptide: Connecting peptide linking A and B chains of pro-insulin, removed to form active insulin.

• A and B Peptides: Chains of amino acids forming mature insulin after processing.

• E.coli Expression: Method to produce insulin chains using recombinant DNA technology in bacteria.

• Diabetes: Chronic disorder characterized by high blood glucose due to insulin deficiency or resistance.

• Allergic Reaction: Immune response against foreign proteins, potentially triggered by animal-derived insulin.

• Disulphide Bond: Covalent linkage joining A and B chains to form functional insulin.

• Recombinant DNA Technology: Technique to insert insulin gene into bacterial plasmid for mass production.

• Mature Insulin: Functional hormone after removal of C-peptide and proper chain folding.

• Exogenous Insulin: Insulin administered from external sources to treat diabetes.

Lead Question - 2022:
Statements related to human Insulin are given below. Which statement(s) is/are correct about genetically engineered Insulin?
(a) Pro-hormone insulin contain extra stretch of C-peptide
(b) A-peptide and B-peptide chains of insulin were produced separately in E.coli, extracted and combined by creating disulphide bond between them
(c) Insulin used for treating Diabetes was extracted from Cattles and Pigs
(d) Pro-hormone Insulin needs to be processed for converting into a mature and functional hormone
(e) Some patients develop allergic reactions to the foreign insulin
Choose the most appropriate answer from the options given below:
(1) (b) only
(2) (c) and (d) only
(3) (c), (d) and (e) only
(4) (a), (b) and (d) only

Explanation: The correct answer is (4) (a), (b) and (d) only. Genetically engineered insulin involves pro-hormone insulin containing extra C-peptide (a), production of A and B chains separately in E.coli and combining them via disulphide bonds (b), and processing pro-insulin into mature functional insulin (d). Animal insulin extraction (c) and allergic reactions (e) are not relevant here.

1. Single Correct Answer MCQ:
Which component is removed from pro-insulin to produce mature insulin?
(a) A-chain
(b) B-chain
(c) C-peptide
(d) Disulphide bond

Explanation: The correct answer is (c) C-peptide. Pro-insulin contains an extra C-peptide connecting the A and B chains. During maturation, the C-peptide is cleaved enzymatically to form functional insulin. The A and B chains remain connected via disulphide bonds to ensure proper biological activity.

2. Single Correct Answer MCQ:
Which microorganism is commonly used to produce insulin chains via recombinant DNA technology?
(a) Saccharomyces cerevisiae
(b) E.coli
(c) Bacillus subtilis
(d) Streptococcus pyogenes

Explanation: The correct answer is (b) E.coli. The A and B peptide chains of insulin are produced separately in E.coli using recombinant DNA technology. Bacteria are preferred due to their rapid growth and ease of genetic manipulation, which allows mass production of insulin efficiently and cost-effectively.

3. Single Correct Answer MCQ:
Disulphide bonds in insulin connect which of the following?
(a) Two C-peptides
(b) A and B chains
(c) Insulin and glucagon
(d) Beta cells

Explanation: The correct answer is (b) A and B chains. Disulphide bonds are covalent linkages that connect the A and B chains of insulin, stabilizing the molecule and maintaining its biological activity. Proper formation of these bonds is essential for insulin to function effectively in glucose regulation.

4. Single Correct Answer MCQ:
What is the primary purpose of producing insulin using recombinant DNA technology?
(a) Reduce allergic reactions
(b) Mass production of human insulin
(c) Extract insulin from animals
(d) Destroy C-peptide

Explanation: The correct answer is (b) Mass production of human insulin. Recombinant DNA technology allows the insertion of human insulin gene into E.coli for efficient production. This method avoids dependence on animal sources, ensures consistency, reduces contamination risks, and provides sufficient insulin to meet global therapeutic demand for diabetic patients.

5. Single Correct Answer MCQ:
Which chain of insulin contains the active site interacting with insulin receptor?
(a) A-chain
(b) B-chain
(c) C-peptide
(d) None

Explanation: The correct answer is (b) B-chain. The B-chain of insulin is primarily responsible for receptor binding and activating insulin signaling pathways. While the A-chain stabilizes the structure via disulphide bonds, the B-chain interacts directly with the insulin receptor, regulating glucose uptake in target tissues.

6. Single Correct Answer MCQ:
Which of the following statements about pro-hormone insulin is correct?
(a) It is biologically active
(b) Contains extra C-peptide
(c) Cannot be processed
(d) Found only in animals

Explanation: The correct answer is (b) Contains extra C-peptide. Pro-hormone insulin is an inactive precursor containing the C-peptide linking the A and B chains. It requires enzymatic processing to remove C-peptide and generate mature, biologically active insulin capable of regulating blood glucose effectively.

7. Assertion-Reason MCQ:
Assertion (A): Recombinant insulin reduces allergic reactions compared to animal insulin.
Reason (R): It is structurally identical to human insulin.
(a) A is true, R is true and R is correct explanation of A
(b) A is true, R is true but R is not correct explanation of A
(c) A is true, R is false
(d) A is false, R is true

Explanation: The correct answer is (a) A is true, R is true and R is correct explanation of A. Recombinant insulin is human-identical and does not carry foreign proteins from pigs or cattle, which reduces immune-mediated allergic reactions in patients, providing safer and more consistent therapy.

8. Matching Type MCQ:
Match the following:
Column I: 1. A-chain 2. B-chain 3. C-peptide
Column II: a. Connects A and B chains b. Receptor binding c. Stabilizes structure
(a) 1-c, 2-b, 3-a
(b) 1-b, 2-c, 3-a
(c) 1-a, 2-b, 3-c
(d) 1-c, 2-a, 3-b

Explanation: The correct answer is (a) 1-c, 2-b, 3-a. The A-chain stabilizes insulin structure, the B-chain binds to the receptor, and the C-peptide connects A and B chains in pro-insulin, which is removed to produce mature, active insulin capable of regulating blood glucose levels efficiently.

9. Fill in the Blanks MCQ:
_________ technology allows production of human insulin by inserting its gene into bacterial plasmids.
(a) Hybridoma
(b) Recombinant DNA
(c) CRISPR
(d) Monoclonal

Explanation: The correct answer is (b) Recombinant DNA. Recombinant DNA technology inserts the human insulin gene into bacterial plasmids, enabling E.coli to produce A and B chains separately. After disulphide bond formation, functional human insulin is obtained, facilitating large-scale production for diabetes treatment.

10. Choose the Correct Statements MCQ:
Which of the following statements about genetically engineered insulin is/are correct?
(a) Produced using E.coli
(b) Contains C-peptide in mature form
(c) Mass production possible
(d) Less immunogenic than animal insulin

Explanation: The correct answer is (a), (c), (d). Recombinant insulin is produced in E.coli (a), allows mass production (c), and is less immunogenic than animal insulin (d). The C-peptide is removed to form mature insulin, so (b) is incorrect. This ensures functional, human-identical insulin for therapeutic use.

Copic: Recombinant DNA Technology
Subtopic: Restriction Endonucleases and DNA Manipulation

Keyword Definitions:
• Restriction endonucleases: Enzymes that cut DNA at specific sequences.
• Palindromic sequence: DNA sequence that reads the same 5' to 3' on both strands.
• Sticky ends: Overhanging single-stranded ends produced by restriction enzymes.
• Blunt ends: DNA ends with no overhang after cleavage.
• DNA ligase: Enzyme that joins DNA fragments by forming phosphodiester bonds.
• Recombinant DNA: DNA formed by joining fragments from different sources.
• Vector: DNA molecule used to carry foreign DNA into a host.
• Cloning: Process of producing identical copies of DNA or organisms.
• Transformation: Introduction of foreign DNA into a host cell.
• Genome editing: Techniques to modify DNA sequences in organisms.

Lead Question (2022):
Given below are two statements:
Statement I: Restriction endonucleases recognise specific sequence to cut DNA known as palindromic nucleotide sequence.
Statement II: Restriction endonucleases cut the DNA strand a little away from the centre of the palindromic site.
In the light of the above statements, choose the most appropriate answer from the options given below:
(1) Both Statement I and Statement II are incorrect
(2) Statement I is correct but Statement II is incorrect
(3) Statement I is incorrect but Statement II is correct
(4) Both Statement I and Statement II are correct

Explanation: The correct answer is (2). Statement I is correct because restriction endonucleases specifically recognise palindromic DNA sequences to cut. Statement II is incorrect as many restriction enzymes cut exactly at the palindromic site producing either sticky or blunt ends, not necessarily away from the center.

Guessed MCQs:

1. Single Correct Answer:
Which type of ends do restriction enzymes produce when they cut DNA?
(a) Sticky ends
(b) Blunt ends
(c) Both sticky and blunt ends
(d) None
Explanation: The correct answer is (c). Restriction enzymes can produce sticky ends with overhangs or blunt ends with no overhang, depending on the enzyme, facilitating ligation of DNA fragments in recombinant DNA technology.

2. Single Correct Answer:
Which enzyme joins DNA fragments in recombinant DNA technology?
(a) Restriction endonuclease
(b) DNA ligase
(c) RNA polymerase
(d) DNA polymerase
Explanation: The correct answer is (b). DNA ligase catalyzes the formation of phosphodiester bonds between DNA fragments, allowing construction of recombinant DNA molecules for cloning or genetic engineering.

3. Single Correct Answer:
A palindromic sequence in DNA:
(a) Is the same in 5’→3’ direction on both strands
(b) Has unequal strands
(c) Codes for proteins
(d) Is always single-stranded
Explanation: The correct answer is (a). Palindromic sequences read identically on both complementary DNA strands in 5’→3’ direction, making them recognition sites for restriction endonucleases.

4. Assertion-Reason MCQ:
Assertion (A): Restriction enzymes are essential for recombinant DNA technology.
Reason (R): They cut DNA at specific sequences, enabling insertion into vectors.
(a) Both A and R are correct and R explains A
(b) Both A and R are correct but R does not explain A
(c) A is correct, R is incorrect
(d) A is incorrect, R is correct
Explanation: The correct answer is (a). Restriction enzymes precisely cut DNA, allowing foreign DNA fragments to be inserted into vectors, which is central to recombinant DNA technology.

5. Single Correct Answer:
Which of the following acts as a carrier for foreign DNA in cloning?
(a) Vector
(b) Restriction enzyme
(c) DNA ligase
(d) Ribosome
Explanation: The correct answer is (a). Vectors such as plasmids or viral genomes carry foreign DNA into host cells for replication and expression in genetic engineering.

6. Single Correct Answer:
Which type of restriction enzyme cuts at a defined distance from recognition site?
(a) Type I
(b) Type II
(c) Type III
(d) Type IV
Explanation: The correct answer is (c). Type III restriction enzymes cut DNA at a short distance from their recognition site, unlike Type II which cut precisely at palindromic sequences.

7. Matching Type:
Match the enzyme with its function:
Column A
1. DNA ligase
2. Restriction endonuclease
3. DNA polymerase
4. Reverse transcriptase
Column B
A. Synthesizes DNA from RNA
B. Cuts DNA at specific sites
C. Joins DNA fragments
D. Synthesizes new DNA strand
Options:
(a) 1-C, 2-B, 3-D, 4-A
(b) 1-B, 2-C, 3-A, 4-D
(c) 1-D, 2-A, 3-C, 4-B
(d) 1-A, 2-D, 3-B, 4-C
Explanation: The correct answer is (a). DNA ligase joins fragments (1-C), restriction endonuclease cuts DNA (2-B), DNA polymerase synthesizes DNA (3-D), reverse transcriptase synthesizes DNA from RNA (4-A).

8. Fill in the Blanks:
Restriction endonucleases recognise ______ sequences and produce ______ ends.
(a) Random, sticky
(b) Palindromic, sticky or blunt
(c) Coding, blunt
(d) Palindromic, only blunt
Explanation: The correct answer is (b). Restriction enzymes specifically recognise palindromic sequences and generate either sticky or blunt ends depending on the enzyme.

9. Single Correct Answer:
Which is the main application of restriction enzymes?
(a) Protein synthesis
(b) Recombinant DNA technology
(c) DNA replication
(d) RNA transcription
Explanation: The correct answer is (b). Restriction enzymes are crucial in recombinant DNA technology for cutting and manipulating DNA to create novel genetic constructs.

10. Choose the correct statements:
(a) Restriction enzymes cut at specific sequences
(b) Palindromic sequences are recognition sites
(c) All restriction enzymes produce sticky ends only
(d) DNA ligase joins DNA fragments
Explanation: The correct answer is (a), (b), and (d). Statement (c) is incorrect because restriction enzymes can produce sticky or blunt ends depending on the enzyme.

Topic: Gene Therapy
Subtopic: Adenosine Deaminase (ADA) Deficiency Treatment

Keyword Definitions:
• Gene therapy: Treatment that introduces functional genes into cells to correct genetic disorders.
• Adenosine Deaminase (ADA): Enzyme crucial for purine metabolism, deficiency leads to severe combined immunodeficiency (SCID).
• Lymphocytes: White blood cells responsible for immune responses.
• Retroviral vector: Virus used to deliver genetic material into cells.
• Genetically engineered cells: Cells modified to carry desired genes.
• Immortal cells: Cells capable of indefinite division.
• Ex vivo therapy: Cells are modified outside the body and reintroduced.
• Infusion: Introduction of fluids or cells into the bloodstream.
• Purine metabolism: Biochemical pathway involving adenine and guanine nucleotides.
• SCID: Severe Combined Immunodeficiency, a disease of defective immune function.

Lead Question (2022):
In gene therapy of Adenosine Deaminase (ADA) deficiency, the patient requires periodic infusion of genetically engineered lymphocytes because:
(1) Gene isolated from marrow cells producing ADA is introduced into cells at embryonic stages
(2) Lymphocytes from patient’s blood are grown in culture, outside the body.
(3) Genetically engineered lymphocytes are not immortal cells.
(4) Retroviral vector is introduced into these lymphocytes.

Explanation: The correct answer is (3). Genetically engineered lymphocytes in ADA gene therapy are modified ex vivo to express ADA but are not immortal. These cells have limited lifespan, so periodic infusions are required to maintain adequate enzyme levels and restore immune function, ensuring continuous protection against infections in ADA-deficient patients.

Guessed MCQs:

1. Single Correct Answer:
Which enzyme deficiency causes severe combined immunodeficiency in ADA gene therapy?
(a) Adenosine Deaminase
(b) DNA Polymerase
(c) RNAse H
(d) Reverse Transcriptase
Explanation: The correct answer is (a). ADA deficiency impairs purine metabolism in lymphocytes, leading to SCID. Gene therapy restores enzyme function to improve immune response.

2. Single Correct Answer:
What type of vector is commonly used in ADA gene therapy?
(a) Retroviral vector
(b) Plasmid vector
(c) Adenoviral vector
(d) Bacteriophage vector
Explanation: The correct answer is (a). Retroviral vectors integrate the ADA gene into lymphocyte DNA, enabling transient or long-term enzyme expression.

3. Single Correct Answer:
Ex vivo gene therapy refers to:
(a) Modifying cells inside the body
(b) Modifying cells outside the body
(c) Modifying embryonic cells only
(d) Injecting naked DNA directly
Explanation: The correct answer is (b). In ex vivo therapy, patient cells are removed, genetically engineered outside the body, and reintroduced to restore normal function.

4. Assertion-Reason MCQ:
Assertion (A): ADA-deficient patients require repeated cell infusion.
Reason (R): Genetically engineered lymphocytes are not immortal.
(a) Both A and R are correct and R explains A
(b) Both A and R are correct but R does not explain A
(c) A is correct, R is incorrect
(d) A is incorrect, R is correct
Explanation: The correct answer is (a). ADA-deficient lymphocytes modified ex vivo do not divide indefinitely, so repeated infusions are essential to maintain immune function.

5. Single Correct Answer:
Which cell type is primarily modified in ADA gene therapy?
(a) Neutrophils
(b) Lymphocytes
(c) Erythrocytes
(d) Platelets
Explanation: The correct answer is (b). Lymphocytes are the target cells for ADA gene therapy since their proper function is critical for adaptive immunity.

6. Single Correct Answer:
Why is ADA gene therapy preferred over bone marrow transplantation in some patients?
(a) Less risk of immune rejection
(b) Requires general anesthesia
(c) Permanent solution
(d) Not effective in children
Explanation: The correct answer is (a). ADA gene therapy uses patient’s own cells, reducing graft-versus-host reactions and immune rejection compared to donor transplantation.

7. Matching Type:
Match Column A (Therapy Components) with Column B (Function):
Column A
1. Retroviral vector
2. Lymphocytes
3. Ex vivo culture
4. ADA enzyme
Column B
A. Delivery of gene into cells
B. Target cells for therapy
C. Site of cell modification
D. Correct metabolic defect
Options:
(a) 1-A, 2-B, 3-C, 4-D
(b) 1-B, 2-A, 3-D, 4-C
(c) 1-A, 2-C, 3-B, 4-D
(d) 1-D, 2-B, 3-A, 4-C
Explanation: The correct answer is (a). Retroviral vector delivers ADA gene (1-A), lymphocytes are targets (2-B), ex vivo culture modifies cells (3-C), ADA enzyme corrects deficiency (4-D).

8. Fill in the Blanks:
In ADA gene therapy, patient’s ______ are genetically engineered ex vivo and reinfused periodically to restore ______.
(a) Lymphocytes, immune function
(b) Neutrophils, oxygen transport
(c) Platelets, clotting function
(d) Red blood cells, hemoglobin
Explanation: The correct answer is (a). Lymphocytes are modified outside the body and reinfused to restore immune responses compromised by ADA deficiency.

9. Single Correct Answer:
Which of the following limits the duration of ADA gene therapy effectiveness?
(a) Short-lived lymphocytes
(b) Vector toxicity
(c) Patient age
(d) Enzyme overdose
Explanation: The correct answer is (a). Genetically modified lymphocytes have limited lifespan, requiring repeated infusions to maintain adequate ADA enzyme activity in patients.

10. Choose the correct statements:
(a) ADA gene therapy uses patient’s own cells
(b) Ex vivo culture allows gene insertion
(c) Genetically engineered lymphocytes are immortal
(d) Periodic infusion is required to maintain function
Explanation: The correct answer is (a), (b), and (d). Patient’s lymphocytes are modified ex vivo with ADA gene and reinfused; they are not immortal, so periodic infusion is essential.

Topic: DNA Structure and Function
Subtopic: Restriction Enzymes and Palindromic Sequences

Keyword Definitions:
• DNA: Deoxyribonucleic acid, carrier of genetic information.
• Restriction Enzyme: Protein that cuts DNA at specific sequences.
• Palindromic Sequence: DNA sequence that reads the same 5’ to 3’ on both strands.
• Base Pair: Complementary nucleotides A-T and G-C forming DNA double helix.
• Recognition Site: Specific DNA sequence recognized and cleaved by a restriction enzyme.
• EcoRI: Common restriction enzyme that cuts 5’ GAATTC 3’.
• Double-stranded DNA: Two complementary DNA strands forming a helix.
• Cleavage: Cutting of DNA by enzymes.
• Overhang: Single-stranded ends produced after cleavage.
• Sticky Ends: Complementary single-stranded DNA overhangs generated by restriction enzyme cuts.

Lead Question (2022):
In the following palindromic base sequences of DNA, which one can be cut easily by a particular restriction enzyme?
Options:
1. 5’ GAATTC 3’ ; 3’ CTTAAG 5’
2. 5’ CTCAGT 3’ ; 3’ GAGTCA 5’
3. 5’ GTATTC 3’ ; 3’ CATAAG 5’
4. 5’ GATACT 3’ ; 3’ CTATGA 5’

Explanation: The correct answer is 1. 5’ GAATTC 3’ ; 3’ CTTAAG 5’ is a palindromic sequence recognized and cleaved efficiently by EcoRI restriction enzyme. Palindromic sequences are ideal recognition sites for restriction enzymes, allowing specific cleavage and generation of sticky ends, crucial for molecular cloning and recombinant DNA technology.

Guessed MCQs:

1. Which enzyme cuts DNA at specific palindromic sequences?
Options:
(a) DNA polymerase
(b) Restriction enzyme
(c) Ligase
(d) RNA polymerase
Explanation: The correct answer is (b) Restriction enzyme. These enzymes recognize specific palindromic sequences in double-stranded DNA and cleave them precisely. They are widely used in genetic engineering and molecular cloning to create fragments for analysis or recombination.

2. EcoRI recognizes which DNA sequence?
Options:
(a) 5’ GAATTC 3’
(b) 5’ CTCAGT 3’
(c) 5’ GTATTC 3’
(d) 5’ GATACT 3’
Explanation: The correct answer is (a) 5’ GAATTC 3’. EcoRI is a type II restriction enzyme that cuts at this palindromic site, producing sticky ends. Other sequences are not recognized by EcoRI, illustrating the specificity of restriction enzymes for their recognition sites.

3. Assertion-Reason MCQ:
Assertion (A): Palindromic sequences are targets for restriction enzymes.
Reason (R): They read the same 5’ to 3’ on complementary strands.
Options:
(a) Both A and R are true, R explains A
(b) Both A and R are true, R does not explain A
(c) A is true, R is false
(d) A is false, R is true
Explanation: The correct answer is (a). Palindromic sequences provide symmetric sites for restriction enzymes to bind and cleave DNA. Complementary strands allow precise recognition and cutting. This symmetry ensures sticky ends are compatible for molecular cloning and recombinant DNA construction.

4. Matching Type MCQ:
Match the restriction enzyme with its recognition site:
List - I List - II
(a) EcoRI (i) 5’ GAATTC 3’
(b) HindIII (ii) 5’ AAGCTT 3’
Options:
1. a-i, b-ii
2. a-ii, b-i
3. a-i, b-i
4. a-ii, b-ii
Explanation: The correct answer is 1. EcoRI recognizes 5’ GAATTC 3’, and HindIII recognizes 5’ AAGCTT 3’. Each enzyme has high specificity for its palindromic site, generating predictable fragments essential for cloning, mapping, and molecular analysis.

5. Which type of ends are generated by EcoRI cleavage?
Options:
(a) Blunt ends
(b) Sticky ends
(c) Hairpin ends
(d) Circular ends
Explanation: The correct answer is (b) Sticky ends. EcoRI cuts DNA asymmetrically within its palindromic site, leaving single-stranded overhangs. Sticky ends facilitate complementary base pairing for ligation in molecular cloning experiments.

6. Single Correct Answer:
Why are palindromic sequences preferred by restriction enzymes?
Options:
(a) They are repetitive
(b) They form hairpins
(c) Symmetric recognition allows precise cleavage
(d) They prevent transcription
Explanation: The correct answer is (c) Symmetric recognition allows precise cleavage. Palindromic sequences read identically on both strands, providing a symmetric site for enzyme binding and cutting, ensuring predictable fragment generation for cloning and molecular biology applications.

7. Fill in the Blanks:
EcoRI produces __________ after cutting DNA.
Options:
(a) Blunt ends
(b) Sticky ends
(c) Mutations
(d) RNA fragments
Explanation: The correct answer is (b) Sticky ends. EcoRI cuts within its palindromic recognition site asymmetrically, leaving single-stranded overhangs known as sticky ends. These ends facilitate annealing with complementary sequences during ligation, crucial for constructing recombinant DNA molecules.

8. Which sequence is NOT palindromic?
Options:
(a) 5’ GAATTC 3’
(b) 5’ GTATTC 3’
(c) 5’ AAGCTT 3’
(d) 5’ GAAGCT 3’
Explanation: The correct answer is (d) 5’ GAAGCT 3’. A palindromic sequence reads identically 5’ to 3’ on complementary strands. This sequence does not fulfill that criterion, unlike GAATTC, GTATTC, and AAGCTT, which are recognized by specific restriction enzymes.

9. Single Correct Answer:
Which enzyme is widely used in recombinant DNA technology for cutting DNA?
Options:
(a) DNA polymerase
(b) Ligase
(c) Restriction enzyme
(d) Helicase
Explanation: The correct answer is (c) Restriction enzyme. Restriction enzymes recognize specific palindromic DNA sequences and cleave them precisely. They are essential tools for cloning, genome mapping, and molecular genetic analysis, enabling construction of recombinant DNA molecules.

10. Choose the correct statements:
(i) Restriction enzymes recognize palindromic sequences
(ii) EcoRI cuts 5’ GAATTC 3’
(iii) Cleavage produces sticky ends
(iv) Only blunt ends are produced
Options:
(a) i, ii, iii
(b) i, ii, iv
(c) ii, iii, iv
(d) i, iii, iv
Explanation: The correct answer is (a) i, ii, iii. Restriction enzymes bind palindromic sequences and cut DNA. EcoRI recognizes 5’ GAATTC 3’, producing sticky ends. Blunt ends are produced by other enzymes, not EcoRI, making sticky ends ideal for recombination and molecular cloning.

Topic: Tools of Recombinant DNA Technology

Subtopic: Transposons and Genetic Engineering Applications

Keyword Definitions:

• Transposons: Segments of DNA that can move from one location to another within the genome, also called jumping genes.

• Gene Silencing: Process by which gene expression is suppressed, preventing the formation of its protein product.

• Gene Sequencing: Technique used to determine the exact sequence of nucleotides in a DNA molecule.

• PCR (Polymerase Chain Reaction): A method to amplify specific DNA sequences in vitro.

• Autoradiography: Technique that uses X-ray film to visualize molecules labeled with radioactive isotopes.

Lead Question (2022):
Transposons can be used during which one of the following?
(1) Gene silencing
(2) Autoradiography
(3) Gene sequencing
(4) Polymerase Chain Reaction

Explanation: Transposons can insert themselves into genes and disrupt their expression, thereby silencing the gene. They are powerful tools in functional genomics to study gene function and regulation. Hence, the correct answer is (1) Gene silencing.

1. Guessed Question:
Which of the following is called “jumping genes”?
(1) Transposons
(2) Plasmids
(3) Exons
(4) Introns

Explanation: Transposons are segments of DNA capable of moving within the genome, often called “jumping genes.” They were first discovered by Barbara McClintock and are used to study gene functions. Hence, the correct answer is (1) Transposons.

2. Guessed Question:
Which scientist discovered transposons?
(1) Watson and Crick
(2) Barbara McClintock
(3) Frederick Sanger
(4) Paul Berg

Explanation: Barbara McClintock discovered transposons in maize during genetic studies. Her discovery earned her the Nobel Prize in Physiology or Medicine in 1983. Hence, the correct answer is (2) Barbara McClintock.

3. Guessed Question:
Which of the following is a function of transposons in genetic studies?
(1) Amplifying DNA sequences
(2) Silencing specific genes
(3) Repairing DNA damage
(4) Synthesizing proteins

Explanation: Transposons can insert into genes, disrupting their function and silencing them. This helps researchers study gene expression and regulation effectively. Hence, the correct answer is (2) Silencing specific genes.

4. Guessed Question:
What is the nature of transposons?
(1) Protein molecules
(2) Mobile DNA segments
(3) RNA fragments
(4) Enzyme molecules

Explanation: Transposons are mobile DNA elements that can move to different genomic positions. They influence genetic variation, gene expression, and evolution. Hence, the correct answer is (2) Mobile DNA segments.

5. Guessed Question:
Which of the following is a common application of transposons in biotechnology?
(1) DNA fingerprinting
(2) Gene silencing
(3) Chromosome counting
(4) Cell staining

Explanation: Transposons are used for gene silencing by insertional mutagenesis, helping scientists understand gene function and regulation. Hence, the correct answer is (2) Gene silencing.

6. Guessed Question:
In which organism were transposons first discovered?
(1) Maize
(2) E. coli
(3) Yeast
(4) Drosophila

Explanation: Transposons were first discovered in maize (corn) by Barbara McClintock. Their discovery explained variations in kernel color and gene movement. Hence, the correct answer is (1) Maize.

7. Assertion-Reason Question:
Assertion (A): Transposons can silence gene expression.
Reason (R): Transposons are mobile elements that integrate into genes and disrupt their transcription.
(1) Both A and R are true, and R explains A.
(2) Both A and R are true, but R does not explain A.
(3) A is true, R is false.
(4) A is false, R is true.

Explanation: Transposons integrate into functional genes, causing disruption of transcription and leading to gene silencing. Both A and R are true, and R correctly explains A. Hence, the correct answer is (1) Both A and R are true, and R explains A.

8. Matching Type Question:
Match the following genetic tools with their primary function:
A. Transposons — (i) Gene silencing
B. Plasmids — (ii) Gene cloning
C. Restriction enzymes — (iii) Cutting DNA
D. PCR — (iv) Amplifying DNA
Options:
(1) A–i, B–ii, C–iii, D–iv
(2) A–ii, B–iii, C–iv, D–i
(3) A–iv, B–i, C–ii, D–iii
(4) A–iii, B–ii, C–i, D–iv

Explanation: Transposons silence genes, plasmids are used for cloning, restriction enzymes cut DNA at specific sites, and PCR amplifies DNA sequences. Hence, the correct answer is (1) A–i, B–ii, C–iii, D–iv.

9. Fill in the Blanks:
Transposons are segments of ______ that can move from one location to another in the ______.
(1) RNA, ribosome
(2) DNA, genome
(3) Protein, nucleus
(4) RNA, cytoplasm

Explanation: Transposons are mobile DNA elements that can shift within the genome, altering gene activity and structure. This property makes them useful tools in genetic engineering. Hence, the correct answer is (2) DNA, genome.

10. Choose the Correct Statements:
(a) Transposons can move within the genome.
(b) They can silence genes.
(c) They are made of RNA.
(d) They are used in genetic engineering.
(1) (a), (b), and (d) only
(2) (b) and (c) only
(3) (a) and (c) only
(4) (a), (b), and (c) only

Explanation: Transposons are mobile DNA segments, can move in the genome, silence genes, and are widely used in genetic engineering for mutagenesis. They are not RNA. Hence, the correct answer is (1) (a), (b), and (d) only.

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Topic: Tools of Recombinant DNA Technology
Subtopic: Gel Electrophoresis Technique

Keyword Definitions:

• Gel Electrophoresis: Technique used to separate DNA, RNA, or proteins based on size and charge.

• Ethidium Bromide: A fluorescent dye that binds to DNA and glows under UV light.

• Elution: The process of extracting separated molecules from a gel after electrophoresis.

• Chromogenic Substrate: A substance that changes color when acted upon by an enzyme, used for visual identification.

Lead Question (2022)
Which one of the following statement is not true regarding gel electrophoresis technique?
(1) The separated DNA fragments are stained by using ethidium bromide
(2) The presence of chromogenic substrate gives blue coloured DNA bands on the gel
(3) Bright orange coloured bands of DNA can be observed in the gel when exposed to UV light
(4) The process of extraction of separated DNA strands from gel is called elution

Explanation:
Chromogenic substrate is not used for staining DNA in gel electrophoresis. DNA fragments are stained using ethidium bromide, which fluoresces orange under UV light. The false statement is about chromogenic substrate producing blue DNA bands. Hence, the correct answer is (2).

1. DNA fragments in agarose gel move towards:
(1) Cathode
(2) Anode
(3) Both electrodes
(4) Remain stationary

Explanation:
DNA is negatively charged due to phosphate groups and moves towards the anode in an electric field during electrophoresis. Hence, the correct answer is (2).

2. Which buffer is used in agarose gel electrophoresis?
(1) Tris-acetate-EDTA
(2) Tris-borate-EDTA
(3) Sodium chloride
(4) Potassium phosphate

Explanation:
Tris-acetate-EDTA (TAE) or Tris-borate-EDTA (TBE) buffers are commonly used to maintain pH and provide ions for current conduction. The correct answer is (1).

3. DNA fragments of which size move fastest through agarose gel?
(1) Large fragments
(2) Small fragments
(3) Medium fragments
(4) All move equally

Explanation:
Smaller DNA fragments encounter less resistance from the gel matrix and migrate faster than larger fragments. Hence, the correct answer is (2).

4. The DNA ladder used in gel electrophoresis helps in:
(1) Measuring voltage
(2) Identifying enzymes
(3) Estimating size of DNA fragments
(4) Increasing current

Explanation:
A DNA ladder contains fragments of known sizes that act as reference markers for estimating the length of sample DNA fragments. Hence, the correct answer is (3).

5. Agarose concentration in gel determines:
(1) Gel color
(2) Resolution of DNA fragments
(3) Electric current
(4) Staining intensity

Explanation:
Higher agarose concentration provides better resolution for small DNA fragments, while lower concentration helps separate larger fragments. Hence, the correct answer is (2).

6. Which factor affects the migration rate of DNA fragments?
(1) Voltage
(2) Gel concentration
(3) DNA size
(4) All of the above

Explanation:
Migration rate depends on DNA size, agarose concentration, and applied voltage. All these influence the speed of DNA movement through the gel. Hence, the correct answer is (4).

7. Assertion-Reason Type:
Assertion (A): DNA fragments move towards the cathode during electrophoresis.
Reason (R): DNA is negatively charged due to phosphate groups.
(1) Both A and R are correct, and R explains A
(2) Both A and R are correct, but R does not explain A
(3) A is false but R is true
(4) Both A and R are false

Explanation:
DNA moves toward the anode, not the cathode, because it is negatively charged. Hence, Assertion is false, but Reason is true. Correct answer is (3).

8. Matching Type:
Match the following:
A. Ethidium bromide — 1. DNA stain
B. Agarose — 2. Gel matrix
C. TAE buffer — 3. Maintains pH
D. DNA ladder — 4. Molecular marker

(1) A–1, B–2, C–3, D–4
(2) A–4, B–1, C–2, D–3
(3) A–3, B–4, C–1, D–2
(4) A–2, B–3, C–4, D–1

Explanation:
Ethidium bromide stains DNA, agarose acts as the gel matrix, buffer maintains pH, and ladder serves as a molecular marker. Hence, the correct match is (1).

9. Fill in the Blanks:
The process of extracting DNA fragments from agarose gel is called ________.
(1) Isolation
(2) Elution
(3) Purification
(4) Washing

Explanation:
After electrophoresis, the separated DNA bands are cut and extracted from the gel by elution. Hence, the correct answer is (2).

10. Choose the Correct Statements:
(a) DNA migrates toward the cathode.
(b) Ethidium bromide fluoresces under UV light.
(c) Small DNA fragments move slower.
(d) Gel electrophoresis separates DNA by size.

(1) (a) and (b) only
(2) (b) and (d) only
(3) (a), (b), and (c) only
(4) All of the above

Explanation:
Ethidium bromide fluoresces under UV light, and gel electrophoresis separates DNA based on size. DNA moves toward the anode, and smaller fragments move faster. Hence, the correct answer is (2).

Topic: Tools of Recombinant DNA Technology
Subtopic: PCR and Selectable Markers

Keyword Definitions:

• Polymerase Chain Reaction (PCR): A technique to amplify a small segment of DNA, producing millions of copies in a short time using Taq polymerase.

• Selectable Marker: A gene that helps identify transformed cells from non-transformed ones, often providing antibiotic resistance.

• Transformation: The process of introducing foreign DNA into a host cell.

• Vector: A DNA molecule used to carry foreign genetic material into another cell.

• Ampicillin Resistance Gene: A selectable marker that allows only transformed bacteria to survive on ampicillin-containing medium.

Lead Question – 2022

Given below are two statements: one is labelled as Assertion (A) and the other as Reason (R).
Assertion (A): Polymerase chain reaction is used in DNA amplification.
Reason (R): The ampicillin resistant gene is used as a selectable marker to check transformation.
(1) Both (A) and (R) are correct but (R) is not the correct explanation of (A)
(2) (A) is correct but (R) is not correct
(3) (A) is not correct but (R) is correct
(4) Both (A) and (R) are correct and (R) is the correct explanation of (A)

Explanation: PCR amplifies DNA segments using primers and Taq polymerase. Ampicillin resistance is used as a selectable marker during transformation, not during amplification. Hence, both statements are correct but unrelated. Therefore, option (1) is correct. (50 words)

Guessed Questions:

1. The enzyme used in PCR to synthesize new DNA strands is:
(1) DNA ligase
(2) Taq polymerase
(3) Restriction endonuclease
(4) DNA helicase

Explanation: Taq polymerase, derived from Thermus aquaticus, is a heat-stable enzyme used to synthesize new DNA strands during PCR cycles. It withstands high temperatures during denaturation. Hence, option (2) Taq polymerase is correct. (50 words)

2. The selectable marker in pBR322 that provides tetracycline resistance is:
(1) tetR gene
(2) ampR gene
(3) ori gene
(4) HindIII site

Explanation: The pBR322 plasmid contains two selectable marker genes: ampR (ampicillin resistance) and tetR (tetracycline resistance). The tetR gene confers tetracycline resistance and helps in identifying recombinant clones. Hence, option (1) is correct. (50 words)

3. Which of the following is not a step of PCR?
(1) Denaturation
(2) Annealing
(3) Ligation
(4) Extension

Explanation: PCR involves denaturation of DNA strands, annealing of primers, and extension of new strands by Taq polymerase. Ligation is a separate process used to join DNA fragments, not part of PCR. Hence, option (3) is correct. (50 words)

4. Assertion (A): Selectable markers help in identifying transformed cells.
Reason (R): Only cells with recombinant DNA can survive on antibiotic media.
(1) Both A and R are true, and R explains A.
(2) Both A and R are true, but R does not explain A.
(3) A is true, R is false.
(4) A is false, R is true.

Explanation: Both statements are true, and R correctly explains A. Selectable markers such as antibiotic resistance genes allow transformed cells to survive while untransformed ones die. Hence, the correct option is (1). (50 words)

5. Match the following:
A. PCR – (i) DNA amplification
B. Restriction enzyme – (ii) Cutting DNA
C. Ligase – (iii) Joining DNA fragments
D. Vector – (iv) Gene transfer
(1) A–i, B–ii, C–iii, D–iv
(2) A–ii, B–i, C–iii, D–iv
(3) A–iv, B–iii, C–ii, D–i
(4) A–i, B–iii, C–ii, D–iv

Explanation: PCR amplifies DNA (i), restriction enzymes cut DNA (ii), ligases join DNA fragments (iii), and vectors transfer genes (iv). Hence, option (1) A–i, B–ii, C–iii, D–iv is correct. (50 words)

6. Which of the following is used as a vector in genetic engineering?
(1) Taq polymerase
(2) pBR322
(3) DNA ligase
(4) EcoRI

Explanation: pBR322 is a commonly used cloning vector in recombinant DNA technology. It carries selectable markers and restriction sites essential for inserting foreign DNA into host cells. Hence, the correct answer is (2) pBR322. (50 words)

7. Fill in the blank:
The enzyme responsible for joining DNA fragments during recombinant DNA formation is ______.
(1) DNA ligase
(2) DNA helicase
(3) RNA polymerase
(4) Primase

Explanation: DNA ligase catalyzes the formation of phosphodiester bonds between DNA fragments, joining the sugar-phosphate backbones of DNA. This enzyme is essential in creating recombinant DNA molecules. Hence, option (1) DNA ligase is correct. (50 words)

8. Choose the correct statements:
(1) PCR is used for amplifying DNA sequences.
(2) Vectors carry foreign DNA into host cells.
(3) Restriction enzymes cut DNA at specific sites.
(4) All of the above.

Explanation: All statements correctly describe essential tools of recombinant DNA technology. PCR amplifies DNA, vectors transfer genetic material, and restriction enzymes cut DNA precisely. Hence, option (4) all of the above is correct. (50 words)

9. In PCR, primers are:
(1) Small DNA sequences complementary to the target DNA region
(2) Enzymes that join DNA fragments
(3) Proteins that stabilize DNA
(4) RNA molecules acting as catalysts

Explanation: Primers are short, single-stranded DNA sequences that initiate DNA synthesis by binding to complementary regions on target DNA. They define the start and end points of amplification. Hence, the correct answer is (1). (50 words)

10. During PCR, the denaturation step occurs at:
(1) 37°C
(2) 72°C
(3) 94°C
(4) 55°C

Explanation: Denaturation separates the two DNA strands by breaking hydrogen bonds at around 94°C. This allows primers to bind during the annealing step. Hence, option (3) 94°C is correct. (50 words)

Copic: Immunology and Diagnostics

Subtopic: Molecular Diagnostic Techniques

Keyword Definitions:

• ELISA Enzyme-Linked Immunosorbent Assay used for detecting antigens or antibodies in patient samples with high sensitivity.

• Southern blotting A DNA detection technique used mainly in genetic studies and DNA fingerprinting.

• Western blotting Protein analysis method used for detecting specific proteins with antibodies.

• Hybridization technique Method of detecting nucleic acid sequences by base pairing with complementary probes.

• Pathophysiology Study of functional changes in the body caused by disease processes.

Lead Question - 2021

For effective treatment of the disease, early diagnosis and understanding its pathophysiology is very important. Which of the following molecular diagnostic techniques is very useful for early detection?
(1) Southern Blotting Technique
(2) ELISA Technique
(3) Hybridization Technique
(4) Western Blotting Technique

Explanation: Early diagnosis of infections depends on detecting small amounts of antigens or antibodies. ELISA is widely used because of its high sensitivity, speed, and specificity. Southern blotting detects DNA, Western blotting proteins, while hybridization is more research-based. Thus, ELISA technique is most effective. Correct answer is option (2).

1. Which diagnostic test is commonly used to confirm HIV infection?
(1) ELISA
(2) MRI
(3) X-ray
(4) CT Scan

Explanation: HIV detection is initially screened by ELISA to identify antibodies against HIV. Confirmation is then done by Western blotting for specificity. Imaging techniques like MRI, X-ray, and CT scans cannot detect HIV. Thus, the correct answer is ELISA.

2. The principle of ELISA is based on:
(1) Antigen-antibody interaction
(2) Enzyme-substrate binding
(3) DNA replication
(4) RNA transcription

Explanation: ELISA functions by detecting antigen-antibody interactions. An enzyme-linked antibody binds to the antigen, and the reaction with a substrate produces a detectable color change. Thus, the correct answer is antigen-antibody interaction.

3. Southern blotting is mainly used for:
(1) RNA detection
(2) Protein detection
(3) DNA detection
(4) Antibody detection

Explanation: Southern blotting involves transferring DNA fragments onto a membrane and hybridizing with complementary probes for detection. It is not used for RNA, protein, or antibody detection. Therefore, the correct answer is DNA detection.

4. Assertion (A): Western blotting is used for protein detection.
Reason (R): It separates proteins by SDS-PAGE and identifies them with specific antibodies.
(1) Both A and R are true, and R explains A
(2) Both A and R are true, but R does not explain A
(3) A is true, R is false
(4) A is false, R is true

Explanation: Western blotting involves separation of proteins by electrophoresis followed by transfer to a membrane and detection with antibodies. Both Assertion and Reason are true, and Reason explains Assertion. Thus, the correct answer is option (1).

5. Match the following:
A. ELISA - i. Antigen-antibody interaction
B. Southern blot - ii. DNA detection
C. Western blot - iii. Protein detection
D. Northern blot - iv. RNA detection
Options:
(1) A-i, B-ii, C-iii, D-iv
(2) A-ii, B-iii, C-i, D-iv
(3) A-iii, B-i, C-iv, D-ii
(4) A-iv, B-ii, C-i, D-iii

Explanation: ELISA is based on antigen-antibody interaction, Southern blot detects DNA, Western blot detects proteins, and Northern blot is used for RNA. Hence, the correct matching is option (1).

6. Which of the following is not a blotting technique?
(1) Southern blot
(2) Northern blot
(3) Eastern blot
(4) Western blot

Explanation: Southern blot detects DNA, Northern blot RNA, Western blot proteins. Eastern blot is not a standard recognized blotting technique. Thus, the correct answer is Eastern blot.

7. Fill in the blank:
__________ technique is most commonly used for detection of viral antigens in blood samples.
(1) ELISA
(2) PCR
(3) CT Scan
(4) Ultrasound

Explanation: ELISA is the most commonly used technique for detecting viral antigens or antibodies in patient blood samples. PCR detects nucleic acids but not directly antigens. Imaging techniques cannot detect viruses. Thus, the correct answer is ELISA.

8. Choose the correct statements about PCR:
(1) It amplifies DNA sequences
(2) It uses primers
(3) It involves Taq polymerase
(4) All of these

Explanation: Polymerase Chain Reaction (PCR) is used to amplify DNA sequences exponentially. It requires primers and thermostable Taq polymerase. All given statements are correct. Therefore, the correct answer is all of these.

9. Which molecular diagnostic method is best for studying gene expression?
(1) Western blot
(2) Northern blot
(3) Southern blot
(4) ELISA

Explanation: Northern blotting is used to study RNA levels, which represent gene expression in cells. Southern blot detects DNA, Western blot proteins, and ELISA antigen-antibody interactions. Thus, the correct answer is Northern blot.

10. In ELISA test, the substrate is used to:
(1) Bind antigen
(2) Produce color change
(3) Denature proteins
(4) Amplify DNA

Explanation: In ELISA, the enzyme-conjugated antibody reacts with a specific substrate to produce a measurable color change. This allows detection of the antigen-antibody complex. Therefore, the correct answer is produce color change.

Topic: Tools of Recombinant DNA Technology

Subtopic: Restriction Enzymes

• Endonucleases: Enzymes that cut DNA at specific internal sites.

• Recognition sequence: Short DNA sequence identified by restriction enzymes for cleavage.

• Palindromic sequence: DNA sequence that reads the same in 5’ to 3’ direction on both strands.

• Okazaki fragments: Short DNA fragments formed during lagging strand synthesis.

• Poly(A) tail: Stretch of adenine bases added to mRNA for stability.

• Degenerate primer: Primer designed to hybridize with multiple related DNA sequences.

• DNA cleavage: Process of cutting DNA molecules by restriction enzymes.

• Recombinant DNA: DNA formed by joining DNA from different sources.

• Restriction site: Specific DNA location recognized by restriction endonucleases.

• Genetic engineering: Manipulation of an organism's DNA to achieve desired traits.

• Molecular scissors: Another name for restriction endonucleases as they cut DNA precisely.

Lead Question - 2021

A specific recognition sequence identified by endonucleases to make cuts at specific positions within the DNA is
Options:
(1) Okazaki sequences
(2) Palindromic Nucleotide sequences
(3) Poly(A) tail sequences
(4) Degenerate primer sequence

Explanation: Restriction endonucleases identify short DNA sequences called palindromic nucleotide sequences, where they make site-specific cuts. These sequences are essential in recombinant DNA technology for creating fragments with sticky or blunt ends. Answer: Palindromic Nucleotide sequences.

1. Single Correct Answer MCQ: Which enzyme acts as molecular scissors in biotechnology?
Options:
A. Ligase
B. DNA Polymerase
C. Restriction Endonuclease
D. RNA Polymerase

Explanation: Restriction endonucleases act as molecular scissors because they cut DNA at specific recognition sites, enabling the creation of recombinant DNA. Answer: Restriction Endonuclease.

2. Single Correct Answer MCQ: Okazaki fragments are formed during:
Options:
A. Leading strand synthesis
B. Lagging strand synthesis
C. mRNA transcription
D. Protein translation

Explanation: Okazaki fragments are short DNA sequences formed on the lagging strand during DNA replication. They are later joined by DNA ligase. Answer: Lagging strand synthesis.

3. Single Correct Answer MCQ: Which structure provides stability to eukaryotic mRNA?
Options:
A. Okazaki fragments
B. Palindromic sequence
C. Degenerate primer
D. Poly(A) tail

Explanation: The Poly(A) tail is added at the 3’ end of eukaryotic mRNA, enhancing stability, transport, and translation efficiency. Answer: Poly(A) tail.

4. Single Correct Answer MCQ: Which type of DNA sequence do restriction endonucleases usually recognize?
Options:
A. Degenerate primers
B. Palindromic sequences
C. Poly(A) tails
D. Okazaki fragments

Explanation: Restriction endonucleases recognize palindromic sequences in DNA and cleave them to generate defined fragments. Answer: Palindromic sequences.

5. Single Correct Answer MCQ: Degenerate primers are useful for:
Options:
A. Recognizing Okazaki fragments
B. Amplifying related DNA sequences
C. Forming Poly(A) tails
D. Acting as endonucleases

Explanation: Degenerate primers contain multiple possible bases at specific positions, allowing them to bind to and amplify related DNA sequences in PCR. Answer: Amplifying related DNA sequences.

6. Single Correct Answer MCQ: Which sequence ensures site-specific cleavage by restriction enzymes?
Options:
A. Palindromic DNA sequence
B. Okazaki fragment
C. Poly(A) tail
D. Degenerate primer

Explanation: Restriction endonucleases cut DNA at palindromic sequences, which are symmetric in base pairing, ensuring site-specific cleavage. Answer: Palindromic DNA sequence.

7. Assertion-Reason MCQ:
Assertion (A): Restriction endonucleases are called molecular scissors.
Reason (R): They cut DNA at specific palindromic sequences.
Options:
A. Both A and R true, R correct explanation
B. Both A and R true, R not correct explanation
C. A true, R false
D. A false, R true

Explanation: Restriction endonucleases act as molecular scissors because they cut DNA precisely at palindromic recognition sequences. Both assertion and reason are true, and R correctly explains A. Answer: Both A and R true, R correct explanation.

8. Matching Type MCQ:
List I: a. Okazaki fragments b. Poly(A) tail c. Restriction endonucleases d. Degenerate primers
List II: i. Molecular scissors ii. RNA stability iii. Lagging strand DNA iv. Multiple DNA targets
Options:
A. a-iii, b-ii, c-i, d-iv
B. a-ii, b-iii, c-iv, d-i
C. a-iv, b-i, c-ii, d-iii
D. a-i, b-iv, c-iii, d-ii

Explanation: Okazaki fragments = lagging strand DNA, Poly(A) tail = RNA stability, Restriction endonucleases = molecular scissors, Degenerate primers = multiple DNA targets. Answer: a-iii, b-ii, c-i, d-iv.

9. Fill in the Blanks MCQ: Short DNA fragments formed on the lagging strand are called _______.
Options:
A. Okazaki fragments
B. Degenerate primers
C. Palindromic sequences
D. Poly(A) tails

Explanation: Short DNA pieces synthesized discontinuously on the lagging strand are called Okazaki fragments. They are later joined by DNA ligase. Answer: Okazaki fragments.

10. Choose the correct statements MCQ:
Options:
A. Restriction endonucleases cut DNA at palindromic sequences
B. Okazaki fragments form on the leading strand
C. Poly(A) tail stabilizes mRNA
D. Degenerate primers can bind to multiple DNA targets
Select:
1. A, B, D
2. A, C, D
3. B, C, D
4. A, C, D

Explanation: Restriction endonucleases cut at palindromic sequences, Poly(A) tail stabilizes mRNA, and degenerate primers bind multiple targets. Okazaki fragments form on the lagging, not leading strand. Correct statements: A, C, D. Answer: Option 4.

Topic: Gene Amplification

Subtopic: Polymerase Chain Reaction (PCR)

• PCR (Polymerase Chain Reaction): A technique to amplify specific DNA sequences using repeated cycles of denaturation, annealing, and extension.

• Denaturation: Step in PCR where double-stranded DNA is separated into single strands using high temperature.

• Annealing: Step in PCR where primers bind to complementary sequences on single-stranded DNA.

• Extension: Step where DNA polymerase synthesizes new DNA strands from primers.

• DNA Polymerase: Enzyme that synthesizes DNA strands using a template strand.

• Primers: Short DNA sequences that guide DNA polymerase to start synthesis at specific sites.

• Gene Amplification: Increasing the number of copies of a specific DNA fragment.

• High Temperature: Required initially in PCR to denature double-stranded DNA for template preparation.

• Ligation: Joining of DNA fragments, not part of PCR amplification.

• Template DNA: DNA sequence to be amplified in PCR.

• Cycling: Repeated sequence of denaturation, annealing, and extension to amplify DNA.

Lead Question - 2021

During the process of gene amplification using PCR, if very high temperature is not maintained in the beginning, then which of the following steps of PCR will be affected first?

(1) Extension
(2) Denaturation
(3) Ligation
(4) Annealing

Explanation: PCR requires high temperature initially for denaturation to separate double-stranded DNA into single strands. Without sufficient heat, the template DNA remains double-stranded, preventing primer binding and subsequent amplification. Other steps, such as annealing and extension, depend on proper denaturation. Answer: Denaturation.

1. Single Correct Answer MCQ: Which PCR step separates double-stranded DNA into single strands?
Options:
A. Denaturation
B. Annealing
C. Extension
D. Ligation

Explanation: Denaturation uses high temperature to separate DNA strands. Annealing and extension occur after denaturation, and ligation is unrelated to PCR. Answer: Denaturation.

2. Single Correct Answer MCQ: During PCR, primers bind to DNA in which step?
Options:
A. Denaturation
B. Annealing
C. Extension
D. Ligation

Explanation: In the annealing step, primers attach to complementary single-stranded DNA after denaturation, guiding DNA polymerase for synthesis. Answer: Annealing.

3. Single Correct Answer MCQ: DNA polymerase adds nucleotides in which PCR step?
Options:
A. Denaturation
B. Annealing
C. Extension
D. Ligation

Explanation: Extension is catalyzed by DNA polymerase to synthesize new DNA strands using primers as starting points. Denaturation and annealing precede extension. Answer: Extension.

4. Single Correct Answer MCQ: What temperature is critical to start PCR?
Options:
A. Low temperature
B. Moderate temperature
C. Very high temperature
D. Room temperature

Explanation: Very high temperature is required initially to denature the double-stranded DNA template, ensuring proper primer binding and amplification. Answer: Very high temperature.

5. Single Correct Answer MCQ: Which enzyme is used for synthesizing DNA in PCR?
Options:
A. DNA polymerase
B. Ligase
C. DNase
D. RNA polymerase

Explanation: DNA polymerase catalyzes the addition of nucleotides to the primer during the extension phase, synthesizing new DNA strands. Ligase and DNase are unrelated. Answer: DNA polymerase.

6. Single Correct Answer MCQ: Ligation in PCR is required for:
Options:
A. Joining DNA fragments
B. Separating strands
C. Primer binding
D. Nucleotide addition

Explanation: Ligation joins DNA fragments but is not part of standard PCR, which relies on denaturation, annealing, and extension. Answer: Joining DNA fragments.

7. Assertion-Reason MCQ:
Assertion (A): Proper high temperature is essential at the start of PCR.
Reason (R): Annealing occurs before denaturation.
Options:
A. Both A and R true, R correct explanation
B. Both A and R true, R not correct explanation
C. A true, R false
D. A false, R true

Explanation: High temperature is necessary for denaturation at the start of PCR. Annealing occurs after denaturation, not before. Assertion true, reason false. Answer: A true, R false.

8. Matching Type MCQ:
List I: a. Denaturation b. Annealing c. Extension d. Ligation
List II: i. DNA strands separate ii. Primers bind iii. DNA synthesis iv. Joining DNA fragments
Options:
A. a-i, b-ii, c-iii, d-iv
B. a-ii, b-i, c-iii, d-iv
C. a-i, b-iii, c-ii, d-iv
D. a-iv, b-i, c-ii, d-iii

Explanation: Denaturation = DNA strands separate, Annealing = primers bind, Extension = DNA synthesis, Ligation = joining fragments. Answer: a-i, b-ii, c-iii, d-iv.

9. Fill in the Blanks MCQ: The first critical step in PCR is _______.
Options:
A. Extension
B. Denaturation
C. Annealing
D. Ligation

Explanation: The denaturation step separates double-stranded DNA into single strands using high temperature, preparing the template for primer binding and amplification. Answer: Denaturation.

10. Choose the correct statements MCQ:
Options:
A. Denaturation requires high temperature
B. Annealing occurs before denaturation
C. DNA polymerase extends primers
D. Ligation is part of PCR
Select:
1. A, B, C
2. A, C
3. B, D
4. All of the above

Explanation: Denaturation needs high temperature (A), and DNA polymerase extends primers (C). Annealing occurs after denaturation, and ligation is not part of PCR. Correct statements: A, C. Answer: A, C.

Subtopic: Insulin Structure and Biosynthesis

• Insulin: Peptide hormone secreted by pancreatic beta cells that regulates blood glucose levels.

• Pro-insulin: Precursor molecule of insulin containing A, B, and C-peptides.

• C-peptide: Connecting peptide cleaved during maturation of insulin; absent in mature insulin.

• A-peptide and B-peptide: Functional chains of insulin connected by disulphide bridges.

• rDNA Technology: Recombinant DNA method used to produce human insulin in bacteria; may include C-peptide depending on method.

• Disulphide Bridges: Covalent bonds connecting cysteine residues of A and B chains for insulin stability.

• Beta Cells: Pancreatic cells producing pro-insulin and insulin.

• Glucose Regulation: Insulin promotes glucose uptake and glycogen synthesis in tissues.

• Peptide Hormones: Hormones composed of amino acid chains like insulin and glucagon.

• Endocrine Secretion: Release of hormones into blood to act on distant target cells.

• Mature Insulin: Functional insulin molecule without C-peptide, active in glucose metabolism.

Lead Question - 2021

With regard to insulin choose correct options:

(a) C-peptide is not present in mature insulin
(b) The insulin produced by rDNA technology has C-peptide
(c) The pro-insulin has C-peptide
(d) A-peptide and B-peptide of insulin are interconnected by disulphide bridges

Options:
1. (b) and (c) only
2. (a), (c) and (d) only
3. (a) and (d) only
4. (b) and (d) only

Explanation: C-peptide is present in pro-insulin but absent in mature insulin. A-peptide and B-peptide are connected by disulphide bridges to form active insulin. Recombinant insulin may lack C-peptide depending on the technology. Correct options reflecting mature insulin structure and biosynthesis are (a), (c) and (d). Answer: (a), (c) and (d) only.

1. Pro-insulin is composed of:
Options:
A. Only A and B chains
B. A, B, and C chains
C. Only C-peptide
D. A and C chains

Explanation: Pro-insulin consists of A-chain, B-chain, and C-peptide. C-peptide connects A and B chains and is removed to form mature insulin. Answer: A, B, and C chains.

2. The role of C-peptide is:
Options:
A. Connect A and B chains in pro-insulin
B. Active in glucose uptake
C. Stabilizes disulphide bonds
D. Hormone receptor binding

Explanation: C-peptide connects the A and B chains in pro-insulin and is cleaved during maturation. It is not directly active in glucose uptake or receptor binding. Answer: Connect A and B chains in pro-insulin.

3. Mature insulin contains:
Options:
A. A, B, C peptides
B. Only A and B peptides
C. Only C-peptide
D. No peptides

Explanation: Mature insulin consists of A and B chains connected by disulphide bridges. The C-peptide is removed during processing and is absent in active insulin. Answer: Only A and B peptides.

4. Disulphide bridges in insulin:
Options:
A. Connect cysteine residues of A and B chains
B. Link C-peptide to A-chain
C. Form alpha helices
D. Bind insulin to receptor

Explanation: Disulphide bridges are covalent bonds connecting cysteine residues of A and B chains, stabilizing insulin’s 3D structure. C-peptide is removed, alpha helices are secondary structures, and bridges do not bind insulin to receptor directly. Answer: Connect cysteine residues of A and B chains.

5. rDNA insulin may differ from natural insulin by:
Options:
A. Presence of C-peptide
B. Absence of A-chain
C. Absence of B-chain
D. No disulphide bonds

Explanation: Recombinant insulin is engineered in bacteria and may retain C-peptide depending on the production method. A and B chains are preserved and disulphide bonds form to create functional insulin. Answer: Presence of C-peptide.

6. Single correct answer: Which peptide is removed to activate insulin?
Options:
A. A-peptide
B. B-peptide
C. C-peptide
D. All peptides

Explanation: During insulin maturation, the C-peptide is cleaved from pro-insulin, leaving the A and B chains connected by disulphide bridges to form active insulin. Answer: C-peptide.

7. Assertion-Reason:
Assertion (A): Disulphide bridges are essential for insulin activity.
Reason (R): They connect A and B chains stabilizing 3D structure.
Options:
A. Both A and R true, R correct explanation
B. Both A and R true, R not correct explanation
C. A true, R false
D. A false, R true

Explanation: The disulphide bridges stabilize insulin’s three-dimensional conformation connecting A and B chains, which is essential for its biological activity. Answer: Both A and R true, R correct explanation.

8. Matching Type:
Column I: 1. Pro-insulin 2. Mature insulin 3. rDNA insulin 4. C-peptide
Column II: A. Active hormone B. Connects A & B chains C. Recombinant preparation D. Precursor with C-peptide
Options:
A. 1-D, 2-A, 3-C, 4-B
B. 1-A, 2-D, 3-B, 4-C
C. 1-C, 2-B, 3-D, 4-A
D. 1-B, 2-C, 3-A, 4-D

Explanation: Correct matching: Pro-insulin – Precursor with C-peptide (D), Mature insulin – Active hormone (A), rDNA insulin – Recombinant preparation (C), C-peptide – Connects A & B chains (B). Answer: 1-D, 2-A, 3-C, 4-B.

9. Fill in the blank: _______ is cleaved from pro-insulin to form active insulin.
Options:
A. A-peptide
B. B-peptide
C. C-peptide
D. Disulphide bridges

Explanation: The C-peptide is removed from pro-insulin, leaving A and B chains linked by disulphide bridges to generate biologically active insulin. Answer: C-peptide.

Topic: Recombinant DNA Technology

Subtopic: Plasmids and Gene Cloning

• Plasmid: Small circular DNA molecule in bacteria, replicates independently of chromosomal DNA.

• pBR322: A commonly used cloning vector with antibiotic resistance genes.

• Restriction Enzyme: Protein cutting DNA at specific sequences for cloning purposes.

• PstI: Restriction enzyme recognizing CTGCAG sequence.

• ampR Gene: Confers ampicillin resistance to bacterial host.

• -Galactoside Production: Enzymatic activity resulting from introduced gene in host.

• Recombinant Plasmid: Plasmid carrying inserted foreign gene.

• Transformed Cells: Bacteria that take up foreign DNA and express its genes.

• Gene Insertion: Integration of foreign DNA into plasmid vector.

• Host Cell Lysis: Breakdown of bacterial cell due to toxic effects or overexpression.

• Novel Protein Expression: Production of a new protein from recombinant DNA.

Lead Question - 2021

Plasmid pBR322 has PstI restriction enzyme site within gene ampR that confers ampicillin resistance. If this enzyme is used for inserting a gene for -galactoside production and the recombinant plasmid is inserted in an E.coli strain:

1. The transformed cells will have the ability to resist ampicillin as well as produce -galactoside.
Options:
A. True
B. False
C. Only -galactoside produced
D. Only ampicillin resistance retained

Explanation: Since the PstI site lies within ampR gene, insertion of the -galactoside gene disrupts ampicillin resistance. The transformed cells can produce -galactoside but will not resist ampicillin. Answer: False.

2. Insertion of foreign gene will lead to lysis of host cell.
Options:
A. True
B. False
C. Only partial lysis
D. Depends on medium

Explanation: Gene insertion in plasmid vectors like pBR322 typically does not cause host cell lysis. Bacterial cells remain viable, allowing expression of inserted genes. Lysis occurs only in cases of toxic gene products or overexpression stress. Answer: False.

3. Recombinant plasmid can produce a novel protein with dual ability.
Options:
A. True
B. False
C. Only native protein
D. Only recombinant protein

Explanation: Since insertion disrupts ampR, the host loses ampicillin resistance. Therefore, the recombinant protein has only the -galactoside activity but not dual function. Answer: False.

4. Recombinant plasmid will not be able to confer ampicillin resistance to the host cell.
Options:
A. True
B. False
C. Only partially
D. Depends on copy number

Explanation: Insertion at the PstI site disrupts the ampR gene. Consequently, the host cannot resist ampicillin, while it can express the inserted -galactoside gene. Answer: True.

5. PstI restriction enzyme recognizes which sequence?
Options:
A. GAATTC
B. CTGCAG
C. GGATCC
D. AAGCTT

Explanation: PstI recognizes the hexanucleotide sequence CTGCAG and cleaves within it. This property is used for targeted gene insertion in vectors like pBR322. Answer: CTGCAG.

6. Host cells that take up recombinant plasmid are called:
Options:
A. Transformed cells
B. Lysed cells
C. Competent cells
D. Mutant cells

Explanation: Bacterial cells that acquire and express foreign DNA are termed transformed cells. They can be selected using markers like antibiotic resistance or reporter gene expression, facilitating identification of successful recombinants. Answer: Transformed cells.

7. Assertion-Reason:
Assertion (A): Inserting a gene at PstI site disrupts ampicillin resistance.
Reason (R): ampR gene is essential for conferring ampicillin resistance.
Options:
A. Both A and R are true, R is correct explanation
B. Both A and R are true, R is not correct explanation
C. A is true, R is false
D. A is false, R is true

Explanation: The ampR gene confers ampicillin resistance. Insertion of foreign DNA at the PstI site interrupts this gene, preventing resistance. Both assertion and reason are correct, and the reason explains the assertion. Answer: Both A and R are true, R is correct explanation.

8. Match the following:
Column I: 1. pBR322 2. ampR 3. PstI 4. Transformed cell
Column II: A. Confers ampicillin resistance B. Vector plasmid C. Host expressing foreign DNA D. Restriction enzyme
Options:
A. 1-B, 2-A, 3-D, 4-C
B. 1-A, 2-B, 3-C, 4-D
C. 1-B, 2-D, 3-A, 4-C
D. 1-C, 2-A, 3-B, 4-D

Explanation: Correct matching: pBR322 – vector plasmid (B), ampR – ampicillin resistance (A), PstI – restriction enzyme (D), Transformed cell – host expressing foreign DNA (C). Answer: 1-B, 2-A, 3-D, 4-C.

9. Fill in the blank: Insertion of foreign gene at ampR site will __________ host’s ampicillin resistance.
Options:
A. Retain
B. Abolish
C. Enhance
D. Partially retain

Explanation: Disruption of ampR by gene insertion abolishes ampicillin resistance. Therefore, transformed cells can express the new gene but cannot survive on ampicillin-containing medium. Answer: Abolish.

10. Choose the correct statements:
1. PstI site lies within ampR gene.
2. Foreign gene insertion disrupts antibiotic resistance.
3. Transformed cells express inserted gene.
Options:
A. 1 and 2 only
B. 2 and 3 only
C. 1 and 3 only
D. 1, 2 and 3

Explanation: All statements are correct. The PstI site is within ampR, foreign gene insertion disrupts ampicillin resistance, and transformed cells can express the inserted -galactoside gene. This forms the basis of selection and expression in recombinant DNA technology. Answer: 1, 2 and 3.

Topic: Polymerase Chain Reaction
Subtopic: Applications of PCR

Keyword Definitions:

• PCR (Polymerase Chain Reaction): Technique to amplify specific DNA segments using primers and DNA polymerase.

• Gene Amplification: Increasing the number of copies of a gene sequence.

• Molecular Diagnosis: Identifying diseases by detecting DNA or RNA sequences.

• Gene Mutation: A permanent change in the DNA sequence of a gene.

• Protein Purification: Laboratory process of isolating proteins from cells or tissues.

Lead Question - 2021

Which of the following is not an application of PCR (Polymerase Chain Reaction)?

(1) Gene amplification
(2) Purification of isolated protein
(3) Detection of gene mutation
(4) Molecular diagnosis

Explanation: PCR is widely used for gene amplification, detection of mutations, and molecular diagnosis of genetic and infectious diseases. However, purification of proteins is not an application of PCR as it deals with DNA amplification, not protein isolation. Therefore, the correct answer is option (2) Purification of isolated protein.

1) Which enzyme is essential in PCR to synthesize new DNA strands?
(1) DNA helicase
(2) Taq polymerase
(3) RNA polymerase
(4) DNA ligase

Explanation: Taq polymerase, a heat-stable DNA polymerase obtained from Thermus aquaticus, is essential for PCR as it withstands high temperatures during denaturation and synthesizes new DNA strands. Hence, the correct answer is option (2) Taq polymerase.

2) Denaturation step in PCR occurs at approximately:
(1) 37°C
(2) 55°C
(3) 72°C
(4) 94°C

Explanation: During PCR, DNA denaturation occurs at around 94°C, where the hydrogen bonds between DNA strands break, forming single-stranded DNA templates for amplification. Therefore, the correct answer is option (4) 94°C.

3) Which of the following is not required in a PCR reaction?
(1) Primers
(2) DNA template
(3) Ribosomes
(4) dNTPs

Explanation: PCR requires primers, DNA template, DNA polymerase, and dNTPs. Ribosomes are involved in protein synthesis, not DNA amplification. Thus, the correct answer is option (3) Ribosomes.

4) In PCR, primers are:
(1) Short DNA sequences
(2) Long RNA molecules
(3) Proteins
(4) Enzymes

Explanation: Primers in PCR are short, single-stranded DNA sequences that provide a starting point for DNA synthesis by binding to complementary DNA regions. Therefore, the correct answer is option (1) Short DNA sequences.

5) Which of the following is an application of PCR?
(1) Cloning genes
(2) Protein folding
(3) Enzyme digestion
(4) Food preservation

Explanation: PCR is used in cloning to amplify specific genes for further studies, genetic engineering, or therapeutic use. It is not used for protein folding, digestion, or food preservation. Correct answer is option (1) Cloning genes.

6) Which of the following is a limitation of PCR?
(1) Requires very small DNA sample
(2) Contamination can lead to false results
(3) Produces results rapidly
(4) Detects DNA mutations efficiently

Explanation: A major limitation of PCR is its sensitivity to contamination, which may lead to false results. Although it requires only small samples and produces rapid results, contamination risk is significant. Correct answer is option (2) Contamination can lead to false results.

7) Assertion (A): PCR can be used in forensic science.
Reason (R): PCR can amplify DNA from minute biological samples such as hair, blood, or saliva.
(1) Both A and R are true, R explains A
(2) Both A and R are true, R does not explain A
(3) A true, R false
(4) A false, R true

Explanation: PCR amplifies DNA from very small biological samples, which makes it useful in forensic science for criminal identification. Both A and R are true, and R correctly explains A. Hence, the correct answer is option (1).

8) Match the following:
(a) Taq polymerase - (i) Heat stable enzyme
(b) Annealing - (ii) Binding of primers
(c) Extension - (iii) DNA synthesis
(d) Denaturation - (iv) Separation of strands
Options:
(1) (a)-(i), (b)-(ii), (c)-(iii), (d)-(iv)
(2) (a)-(ii), (b)-(iii), (c)-(i), (d)-(iv)
(3) (a)-(iii), (b)-(iv), (c)-(ii), (d)-(i)
(4) (a)-(iv), (b)-(i), (c)-(ii), (d)-(iii)

Explanation: Taq polymerase is heat-stable, annealing involves primer binding, extension refers to DNA synthesis, and denaturation separates DNA strands. Thus, the correct matching is option (1).

9) Fill in the blank: The temperature for primer annealing in PCR is generally ______.
(1) 20–25°C
(2) 37–40°C
(3) 50–65°C
(4) 90–95°C

Explanation: During PCR, annealing occurs at 50–65°C, where primers bind to the template DNA strands. This allows Taq polymerase to initiate DNA synthesis during the extension step. Therefore, the correct answer is option (3) 50–65°C.

10) Choose the correct statements:
A. PCR requires thermostable DNA polymerase.
B. PCR involves denaturation, annealing, and extension steps.
C. PCR can detect RNA viruses without conversion.
D. PCR is useful in medical diagnostics.
Options:
(1) A, B, D correct
(2) A, C, D correct
(3) B, C, D correct
(4) A, B, C, D correct

Explanation: PCR requires thermostable polymerase, follows denaturation-annealing-extension steps, and is useful in diagnostics. However, RNA viruses require conversion to complementary DNA (cDNA) before amplification. Hence, A, B, and D are correct. The answer is option (1).

Topic: Recombinant DNA Technology
Subtopic: DNA Purification Techniques

Keyword Definitions:

• DNA (Deoxyribonucleic Acid): The hereditary material in almost all organisms, carrying genetic instructions for development and functioning.

• Histones: Proteins around which DNA winds to form nucleosomes, aiding in chromatin structure and gene regulation.

• Polysaccharides: Carbohydrates composed of long chains of monosaccharide units, often involved in cell wall and energy storage.

• RNA (Ribonucleic Acid): A nucleic acid involved in protein synthesis and gene expression.

• Recombinant DNA Technology: Techniques that combine DNA from different sources to produce new genetic combinations.

• Ethanol Precipitation: A method used to isolate DNA by reducing its solubility in alcohol, causing it to precipitate.

• Chilled Ethanol: Ethanol cooled to low temperatures to efficiently precipitate DNA from aqueous solutions.

• Purification Process: Techniques used to separate and isolate DNA from proteins, RNA, and other cellular components.

Lead Question - 2021

During the purification process for recombinant DNA technology, addition of chilled ethanol precipitates out:
(1) DNA
(2) Histones
(3) Polysaccharides
(4) RNA

Explanation: The correct answer is (1) DNA. In recombinant DNA technology, chilled ethanol is used to precipitate DNA because it reduces DNA solubility. This allows DNA to aggregate and separate from proteins, RNA, and other cellular components, facilitating its purification for further molecular biology applications such as cloning or PCR.

Guessed Questions:

1) Which substance is commonly used to precipitate DNA during extraction?
(1) Chilled ethanol
(2) Warm methanol
(3) Acetone
(4) Chloroform

Explanation: The correct answer is (1) Chilled ethanol. Ethanol at low temperatures decreases the solubility of DNA, causing it to form visible aggregates that can be pelleted by centrifugation. This is a standard step in DNA extraction protocols for isolating pure nucleic acids.

2) Assertion (A): RNA remains soluble in ethanol during DNA precipitation.
Reason (R): RNA has different solubility properties than DNA.
(1) Both A and R true, R explains A
(2) Both A and R true, R does not explain A
(3) A true, R false
(4) A false, R true

Explanation: The correct answer is (1). During ethanol precipitation, DNA precipitates while RNA often remains soluble. This is due to structural differences and solubility properties, allowing selective isolation of DNA while RNA and proteins stay in solution, simplifying purification.

3) Which of the following is used to remove proteins before DNA precipitation?
(1) Phenol-chloroform
(2) Ethanol
(3) Acetic acid
(4) Methanol

Explanation: The correct answer is (1) Phenol-chloroform. This organic solvent mixture denatures and removes proteins from nucleic acid solutions, allowing DNA to be separated during subsequent ethanol precipitation.

4) Fill in the blank: The technique to isolate DNA using alcohol is called _______.
(1) Centrifugation
(2) Gel electrophoresis
(3) Ethanol precipitation
(4) PCR

Explanation: The correct answer is (3) Ethanol precipitation. DNA solubility decreases in alcohol, causing it to aggregate and precipitate out of solution. This step is widely used in molecular biology to purify and concentrate DNA for downstream applications.

5) During DNA extraction, which component binds to the alcohol to facilitate precipitation?
(1) Sugar backbone
(2) Nitrogenous bases
(3) Phosphate groups
(4) Proteins

Explanation: The correct answer is (3) Phosphate groups. DNA's negatively charged phosphate backbone interacts with cations and ethanol, reducing solubility and causing DNA molecules to precipitate, leaving proteins and RNA in solution.

6) Which of the following remains soluble in the aqueous phase after ethanol addition?
(1) DNA
(2) RNA
(3) Both DNA and RNA
(4) None

Explanation: The correct answer is (2) RNA. During ethanol precipitation of DNA, RNA usually remains in the aqueous solution because it has different solubility properties, allowing separation of DNA from RNA during nucleic acid purification.

7) Which of the following cations is often added to facilitate DNA precipitation with ethanol?
(1) Na+
(2) K+
(3) Mg2+
(4) Cl-

Explanation: The correct answer is (1) Na+. Sodium ions neutralize the negatively charged DNA backbone, reducing solubility and promoting precipitation when ethanol is added.

8) Assertion (A): Ethanol precipitation is temperature-dependent.
Reason (R): Chilled ethanol is more effective in precipitating DNA.
(1) Both A and R true, R explains A
(2) Both A and R true, R does not explain A
(3) A true, R false
(4) A false, R true

Explanation: The correct answer is (1). Precipitation efficiency increases at low temperatures because chilled ethanol further reduces DNA solubility, resulting in better aggregation and pellet formation during centrifugation.

9) Matching Type: Match the step with its purpose in DNA purification.
List-I    List-II
(a) Phenol-chloroform extraction    (i) Remove proteins
(b) Ethanol precipitation    (ii) Isolate DNA
(c) RNase treatment    (iii) Remove RNA
(d) Centrifugation    (iv) Separate phases
Select correct answer:
(a) (b) (c) (d)
(1) i, ii, iii, iv
(2) iv, ii, iii, i
(3) i, iv, ii, iii
(4) iii, i, iv, ii

Explanation: The correct answer is (1) i, ii, iii, iv. Phenol-chloroform removes proteins (i), ethanol precipitates DNA (ii), RNase removes RNA (iii), and centrifugation separates aqueous and organic phases (iv), completing the purification process effectively.

10) Choose the correct statements:
a. Chilled ethanol precipitates DNA.
b. RNA is also precipitated by ethanol.
c. Sodium ions enhance DNA precipitation.
d. Phenol-chloroform removes proteins.
(1) a, c, d
(2) a, b, c
(3) b, c, d
(4) a, b, d

Explanation: The correct answer is (1) a, c, d. Chilled ethanol selectively precipitates DNA (a), sodium ions facilitate precipitation (c), and phenol-chloroform removes proteins (d). RNA typically remains in solution, making statement b incorrect.

Topic: Genetic Engineering Techniques
Subtopic: Polymerase Chain Reaction (PCR)

Keyword Definitions:

• Polymerase Chain Reaction (PCR): A molecular technique used to amplify specific DNA sequences in vitro.

• Denaturation: The step in PCR where double-stranded DNA is heated to separate into single strands.

• Annealing: The step in PCR where primers bind to the complementary DNA sequences at lower temperature.

• Extension: The step in PCR where DNA polymerase synthesizes a new DNA strand complementary to the template.

• DNA Polymerase: An enzyme that synthesizes DNA strands during PCR, typically heat-stable Taq polymerase.

• Primers: Short DNA sequences that initiate DNA synthesis in PCR.

• Thermocycler: Equipment used to perform PCR by cycling temperatures for denaturation, annealing, and extension.

• Amplification: Production of multiple copies of a specific DNA fragment using PCR.

Lead Question - 2021

Which of the following is a correct sequence of steps in a PCR (Polymerase Chain Reaction)?
(1) Denaturation, Extension, Annealing
(2) Extension, Denaturation, Annealing
(3) Annealing, Denaturation, Extension
(4) Denaturation, Annealing, Extension

Explanation: The correct answer is (4) Denaturation, Annealing, Extension. In PCR, DNA is first denatured at high temperature to separate strands, then primers anneal to target sequences at lower temperature, and finally DNA polymerase extends the primers to synthesize new strands. This cycle is repeated to amplify DNA exponentially.

Guessed Questions:

1) Which enzyme is commonly used in PCR due to its heat stability?
(1) DNA ligase
(2) Taq polymerase
(3) RNA polymerase
(4) Reverse transcriptase

Explanation: The correct answer is (2) Taq polymerase. This DNA polymerase, isolated from Thermus aquaticus, is heat-stable and can withstand repeated denaturation steps in PCR, making it ideal for synthesizing new DNA strands without denaturing the enzyme.

2) Assertion (A): PCR is used to amplify a specific DNA sequence.
Reason (R): PCR requires primers and DNA polymerase to selectively copy the target region.
(1) Both A and R true, R explains A
(2) Both A and R true, R does not explain A
(3) A true, R false
(4) A false, R true

Explanation: The correct answer is (1). PCR selectively amplifies a target DNA sequence using specific primers and DNA polymerase. Primers determine the start points, and polymerase synthesizes complementary strands, enabling multiple cycles of amplification for large quantities of the specific DNA fragment.

3) Which step of PCR involves primer binding to the target DNA?
(1) Denaturation
(2) Annealing
(3) Extension
(4) Termination

Explanation: The correct answer is (2) Annealing. During this step, the reaction temperature is lowered to allow primers to hybridize with their complementary sequences on the single-stranded DNA, providing starting points for DNA polymerase to synthesize new DNA strands.

4) Fill in the blank: The process of repeated cycles in PCR to increase DNA copy number is called ________.
(1) Denaturation
(2) Amplification
(3) Transcription
(4) Replication

Explanation: The correct answer is (2) Amplification. PCR cycles of denaturation, annealing, and extension lead to exponential production of the target DNA sequence, increasing its quantity significantly from a small starting sample, which is crucial for diagnostics, cloning, and molecular biology studies.

5) Which temperature-dependent step separates double-stranded DNA in PCR?
(1) Denaturation
(2) Annealing
(3) Extension
(4) Ligation

Explanation: The correct answer is (1) Denaturation. High temperature (typically 94–98°C) breaks hydrogen bonds between complementary DNA strands, resulting in single-stranded DNA templates for primer annealing and subsequent synthesis during PCR.

6) Which component provides the template for PCR?
(1) Primers
(2) DNA polymerase
(3) Target DNA
(4) Nucleotides

Explanation: The correct answer is (3) Target DNA. PCR requires a DNA fragment to be amplified. The template DNA contains the specific sequence of interest that primers will bind to, allowing DNA polymerase to synthesize new complementary strands through repeated cycles.

7) Choose the correct statements:
a. PCR can detect genetic mutations.
b. PCR uses repeated temperature cycles.
c. RNA can be directly amplified by standard PCR.
d. Primers determine the target DNA region.
(1) a, b, d
(2) a, c, d
(3) b, c
(4) a, b, c, d

Explanation: The correct answer is (1) a, b, d. PCR detects mutations, uses thermal cycling, and relies on primers for specificity. Statement c is incorrect because RNA requires reverse transcription to cDNA before amplification (RT-PCR). Standard PCR amplifies DNA, not RNA directly.

8) Assertion (A): DNA polymerase synthesizes new DNA strands in PCR.
Reason (R): Taq polymerase remains stable at high denaturation temperatures.
(1) Both A and R true, R explains A
(2) Both A and R true, R does not explain A
(3) A true, R false
(4) A false, R true

Explanation: The correct answer is (1). DNA polymerase synthesizes complementary strands in PCR. Taq polymerase is heat-stable, allowing it to function through repeated denaturation cycles without losing activity, enabling efficient DNA amplification over many PCR cycles.

9) Matching Type: Match the PCR step with its function.
List-I    List-II
(a) Denaturation  (i) DNA synthesis
(b) Annealing  (ii) Strand separation
(c) Extension  (iii) Primer binding
Select correct answer:
(a) (b) (c)
(1) ii, iii, i
(2) i, ii, iii
(3) iii, ii, i
(4) ii, i, iii

Explanation: The correct answer is (1) ii, iii, i. Denaturation separates DNA strands (ii), Annealing allows primers to bind (iii), and Extension synthesizes new DNA (i). This sequence forms the standard PCR cycle repeated multiple times for exponential DNA amplification.

10) Fill in the blank: PCR is widely used in ________ for detecting pathogens, mutations, and forensic analysis.
(1) Biochemistry
(2) Molecular biology
(3) Ecology
(4) Physiology

Topic: Plant Tissue Culture and Genetic Manipulation
Subtopic: Applications of Plant Biotechnology

Keyword Definitions:

• Protoplast Fusion: Fusion of two plant cells without cell walls to combine genetic material.

• Plant Tissue Culture: Growing plant cells or tissues under sterile conditions on a nutrient medium.

• Meristem Culture: Technique of growing meristematic tissue to produce virus-free plants.

• Micropropagation: Rapid cloning of plants using tissue culture techniques.

• Somaclones: Plants derived from somatic cells by tissue culture, genetically identical to the parent.

• Pomato: Hybrid plant obtained by grafting potato and tomato, or via protoplast fusion techniques.

• Virus-Free Plants: Plants regenerated from meristem tissues to eliminate viral infections.

• Totipotency: Ability of a single plant cell to regenerate into a whole plant under suitable conditions.

Lead Question - 2021

Match List - I with List - II.
List - I    List - II
(a) Protoplast fusion  (i) Totipotency
(b) Plant tissue culture  (ii) Pomato
(c) Meristem culture  (iii) Somaclones
(d) Micropropagation  (iv) Virus free plants
Select the correct answer from the options below:
(a) (b) (c) (d)
(1) (ii) (i) (iv) (iii)
(2) (iii) (iv) (i) (ii)
(3) (iv) (iii) (ii) (i)
(4) (iii) (iv) (ii) (i)

Explanation: The correct answer is (1) (ii) (i) (iv) (iii). Protoplast fusion is used to produce hybrids like Pomato, plant tissue culture relies on totipotency to regenerate whole plants, meristem culture generates virus-free plants, and micropropagation produces somaclones, demonstrating various applications of biotechnology in plant improvement.

Guessed Questions:

1) Which technique is primarily used to produce virus-free plants?
(1) Protoplast fusion
(2) Meristem culture
(3) Micropropagation
(4) Callus culture

Explanation: The correct answer is (2) Meristem culture. Meristematic tissues are actively dividing and generally free from viruses. By culturing meristems under sterile conditions, virus-free plants can be regenerated. This technique ensures healthy propagation of plants, especially in crops like banana, potato, and sugarcane, preventing viral disease spread.

2) Assertion (A): Micropropagation produces genetically identical plants.
Reason (R): Somatic cells are totipotent and can develop into a whole plant.
(1) Both A and R true, R explains A
(2) Both A and R true, R does not explain A
(3) A true, R false
(4) A false, R true

Explanation: The correct answer is (1). Micropropagation produces genetically identical plants (somaclones) because somatic cells exhibit totipotency. Under suitable conditions in tissue culture, these cells regenerate into whole plants, maintaining the genetic fidelity of the parent, which allows rapid multiplication of elite or disease-free plant varieties.

3) Which of the following is an example of a hybrid plant produced using protoplast fusion?
(1) Wheat
(2) Pomato
(3) Maize
(4) Rice

Explanation: The correct answer is (2) Pomato. Protoplast fusion combines genetic material from potato and tomato to produce this hybrid. It demonstrates the utility of biotechnological approaches in creating new plant varieties that cannot be obtained through conventional breeding, combining desirable traits from different species.

4) Fill in the blank: The ability of a single plant cell to regenerate into a whole plant is called ________.
(1) Micropropagation
(2) Totipotency
(3) Meristem culture
(4) Somacloning

Explanation: The correct answer is (2) Totipotency. Totipotency allows a single somatic plant cell to develop into an entire plant under appropriate conditions. This property is fundamental to plant tissue culture, micropropagation, and regeneration of plants from callus or protoplasts, enabling mass propagation and biotechnological interventions.

5) Which method is most suitable for rapid multiplication of ornamental plants?
(1) Protoplast fusion
(2) Meristem culture
(3) Micropropagation
(4) Seed propagation

Explanation: The correct answer is (3) Micropropagation. Micropropagation allows large-scale, rapid, and uniform production of plants in a short time. It is especially useful for ornamental and horticultural crops, maintaining desirable traits and eliminating pathogens, making it more efficient than conventional seed propagation or other tissue culture methods.

6) Which of the following results in somaclones?
(1) Meristem culture
(2) Micropropagation
(3) Protoplast fusion
(4) Hybridization

Explanation: The correct answer is (2) Micropropagation. Somaclones are plants regenerated from somatic cells via tissue culture methods. Micropropagation produces multiple genetically identical plants (clones) from explants, ensuring uniformity and preservation of elite genotypes for agricultural and commercial purposes.

7) Choose the correct statements:
a. Protoplast fusion combines genetic material from different species.
b. Meristem culture is used for virus elimination.
c. Micropropagation exploits totipotency.
d. Plant tissue culture cannot regenerate whole plants.
(1) a, b, c
(2) a, c, d
(3) b, d
(4) a, b, c, d

Explanation: The correct answer is (1) a, b, c. Protoplast fusion enables hybrid creation, meristem culture produces virus-free plants, and micropropagation relies on totipotency. Statement d is incorrect because plant tissue culture can regenerate entire plants from cells or tissues, demonstrating its versatility in biotechnology.

8) Assertion (A): Virus-free plants are obtained from meristem cultures.
Reason (R): Meristematic cells are actively dividing and usually free from viruses.
(1) Both A and R true, R explains A
(2) Both A and R true, R does not explain A
(3) A true, R false
(4) A false, R true

Explanation: The correct answer is (1). Meristematic cells are less likely to be infected by viruses due to their high division rate and isolation. Culturing these cells under sterile conditions allows regeneration of virus-free plants, making meristem culture a critical technique for producing healthy, pathogen-free plant material.

9) Which of the following is a

Subtopic: Gene Therapy

Keyword Definitions:

• Gene therapy: Treatment of genetic disorders by introducing, altering, or silencing genes within a patient’s cells.

• Gene targeting: Specific modification of a gene sequence within a genome to achieve desired expression or correction.

• Gene amplification: Increasing the number of copies of a gene for higher expression of its product.

• Molecular diagnosis: Identification of diseases using DNA, RNA, or protein-based techniques.

• Biopiracy: Unauthorized use of biological resources or traditional knowledge for commercial gain.

• Safety testing: Evaluation of a treatment or compound to ensure it does not cause harmful effects.

Lead Question - 2021

When gene targeting involving gene amplification is attempted in an individual's tissue to treat disease, it is known as:
(1) Genetherapy
(2) Molecular diagnosis
(3) Safety testing
(4) Biopiracy

Explanation: The correct answer is (1) Gene therapy. It involves the introduction or correction of genes in target tissues to treat disorders. Gene targeting ensures specific changes, while gene amplification enhances therapeutic gene expression. This approach is used in diseases like hemophilia, immunodeficiency, and certain cancers.

Guessed Questions:

1) Which of the following is an example of in vivo gene therapy?
(1) Direct injection of therapeutic gene into tissues
(2) Gene modification in culture followed by cell transplantation
(3) Prenatal screening for genetic disease
(4) Protein replacement therapy

Explanation: The correct answer is (1) Direct injection of therapeutic gene into tissues. In vivo gene therapy introduces DNA directly into patient cells inside the body. Ex vivo involves culturing and reintroducing modified cells. Screening and protein replacement are diagnostic or supportive methods, not gene therapy.

2) Which genetic disorder has been successfully treated with gene therapy?
(1) Hemophilia
(2) ADA deficiency
(3) Down syndrome
(4) Huntington’s disease

Explanation: The correct answer is (2) ADA deficiency. Adenosine deaminase deficiency causes severe immunodeficiency. Gene therapy using functional ADA gene restores immune function. Hemophilia and Huntington’s are under trials, while Down syndrome is chromosomal, not curable by gene therapy. ADA was one of the first clinical applications.

3) Molecular diagnosis includes:
(1) PCR
(2) ELISA
(3) DNA hybridization
(4) All of the above

Explanation: The correct answer is (4) All of the above. PCR amplifies specific DNA sequences, ELISA detects proteins or antibodies, and DNA hybridization identifies complementary sequences. Together, these techniques form the basis of molecular diagnosis, enabling early detection of diseases, including infections and genetic disorders.

4) Which of the following is an example of biopiracy?
(1) Developing hybrid crops legally
(2) Patenting neem extract without local consent
(3) Conducting safety tests before drug approval
(4) Using CRISPR for gene editing

Explanation: The correct answer is (2) Patenting neem extract without local consent. Biopiracy occurs when corporations exploit indigenous biological resources or traditional knowledge without acknowledgment or compensation. Legal hybrid development, safety tests, and CRISPR applications are scientific practices, not exploitation of community rights.

5) Which technique is used in gene therapy to deliver therapeutic genes?
(1) Bacterial transformation
(2) Viral vectors
(3) Photosynthesis
(4) Gel electrophoresis

Explanation: The correct answer is (2) Viral vectors. Viruses like retroviruses and adenoviruses are engineered to deliver therapeutic DNA into human cells. Bacterial transformation applies to prokaryotes, photosynthesis is unrelated, and gel electrophoresis is a diagnostic tool. Viral vectors ensure efficient and targeted gene delivery in therapy.

6) Which of the following is not a goal of gene therapy?
(1) Correction of defective genes
(2) Treatment of acquired diseases
(3) Cloning of entire human
(4) Production of therapeutic proteins

Explanation: The correct answer is (3) Cloning of entire human. Gene therapy aims at correcting defective genes, treating genetic or acquired diseases, and enhancing therapeutic protein production. Human cloning is ethically unacceptable and not an objective of gene therapy. The therapy remains patient-specific and corrective.

7) Assertion (A): Viral vectors are used in gene therapy.
Reason (R): They can efficiently deliver genes into target cells.
(1) Both A and R are true, R explains A
(2) Both A and R are true, R does not explain A
(3) A is true, R is false
(4) A is false, R is true

Explanation: The correct answer is (1) Both A and R are true, R explains A. Viral vectors naturally infect cells, making them effective carriers of therapeutic DNA. This efficiency explains their extensive use in gene therapy despite challenges like immune responses and insertional mutagenesis risks.

8) Match the following:
A. ADA deficiency - (i) First gene therapy success
B. Retrovirus - (ii) Vector for gene delivery
C. PCR - (iii) Amplification of DNA
D. Biopiracy - (iv) Misuse of biodiversity
Options:
(1) A-i, B-ii, C-iii, D-iv
(2) A-ii, B-iii, C-iv, D-i
(3) A-iii, B-iv, C-i, D-ii
(4) A-iv, B-i, C-ii, D-iii

Explanation: The correct answer is (1) A-i, B-ii, C-iii, D-iv. ADA deficiency was the first disorder treated with gene therapy, retroviruses act as vectors, PCR amplifies DNA, and biopiracy is unauthorized exploitation of biodiversity. Each concept represents a cornerstone in biotechnology and ethics.

9) Fill in the blank: The first successful human gene therapy was performed for __________.
(1) Hemophilia
(2) ADA deficiency
(3) Cystic fibrosis
(4) Sickle-cell anemia

Explanation: The correct answer is (2) ADA deficiency. In 1990, a 4-year-old girl with ADA deficiency underwent gene therapy using functional ADA gene delivered via retroviral vectors. This marked the first success in treating a genetic disorder through gene therapy and opened doors for advanced biomedical research.

10) Choose the correct statements:
A. Gene therapy can be somatic or germline.
B. Biopiracy is ethical usage of traditional knowledge.
C. PCR is a diagnostic technique, not therapy.
D. Viral vectors can cause immune reactions.
Options:
(1) A, C and D
(2) A and B
(3) B, C and D
(4) A, B and D

Explanation: The correct answer is (1) A, C and D. Gene therapy can target somatic or germline cells, though germline is ethically debated. PCR is diagnostic, not therapeutic. Viral vectors may trigger immune responses. Statement B is incorrect as biopiracy represents unethical use of resources.

Topic: Intellectual Property and Bio-resources
Subtopic: Biopiracy and Legal Protection

Keyword Definitions:

• Biopatenting: Legal protection of inventions or processes involving biological material.

• Bioethics: Ethical study of biological research, technology, and applications.

• Bioengineering: Application of engineering principles to biological systems for technology development.

• Biopiracy: Unauthorized use or exploitation of biological resources or traditional knowledge.

• Bio-resources: Plants, animals, microorganisms, or genetic materials used for research or commercial purposes.

• Intellectual property: Legal rights protecting inventions, designs, and knowledge.

• Exploitation: Using biological resources for commercial or personal benefit.

• Traditional knowledge: Indigenous knowledge of bio-resources or medicinal plants.

• Legal protection: Laws and rules preventing misuse or unauthorized access.

• Commercialization: Process of turning bio-resources or knowledge into marketable products.

• Unauthorized use: Exploitation without permission or benefit sharing.

Lead Question - 2020 (COVID Reexam)

The laws and rules to prevent unauthorized exploitation of bio-resources are termed as:

1. Biopatenting
2. Bioethics
3. Bioengineering
4. Biopiracy

Explanation: The correct answer is option 4. Biopiracy refers to the unauthorized use of bio-resources and traditional knowledge without proper consent or benefit sharing. Laws and regulations prevent exploitation by imposing legal protection, ensuring equitable use, and protecting indigenous knowledge from commercial misuse. Biopatenting, bioethics, and bioengineering are related but distinct concepts.

1. Chapter: Biotechnology and Bioethics
Topic: Intellectual Property and Bio-resources
Subtopic: Legal Protection and Ethics

Keyword Definitions:

• Biopiracy: Unauthorized exploitation of bio-resources or knowledge.

• Biopatenting: Legal protection of inventions derived from biological materials.

• Bioethics: Ethical considerations in research and biotechnology.

• Bioengineering: Engineering application to biological systems.

• Bio-resources: Living organisms, genetic material, or products from nature.

• Intellectual property: Rights to protect inventions and innovations.

• Traditional knowledge: Indigenous knowledge regarding plants or natural resources.

• Exploitation: Commercial or unauthorized use of biological resources.

• Legal framework: Laws to regulate bio-resource use.

• Benefit sharing: Equitable distribution of profits from bio-resources.

• Unauthorized use: Using bio-resources without permission.

Q1. Single Correct Answer: Unauthorized use of traditional medicinal plants is termed:
a) Biopatenting
b) Biopiracy
c) Bioethics
d) Bioengineering

Explanation: Using traditional medicinal plants without consent or compensation is biopiracy. Biopatenting protects inventions, bioethics studies ethical issues, and bioengineering applies engineering to biology. Option (b) is correct.

Q2. Single Correct Answer: The legal protection of inventions derived from biological materials is:
a) Bioethics
b) Biopiracy
c) Biopatenting
d) Bioengineering

Explanation: Biopatenting grants legal rights to inventions derived from biological materials, preventing unauthorized use. Bioethics addresses ethical issues, biopiracy is illegal exploitation, and bioengineering involves applying engineering to biology. Option (c) is correct.

Q3. Single Correct Answer: Ethical guidelines for conducting research on bio-resources are termed:
a) Bioethics
b) Biopatenting
c) Biopiracy
d) Bioengineering

Explanation: Bioethics provides ethical guidance in biological research, ensuring responsible use of bio-resources. Biopatenting protects inventions, biopiracy refers to exploitation, and bioengineering applies engineering principles. Option (a) is correct.

Q4. Single Correct Answer: Exploitation of genetic material without consent is an example of:
a) Bioengineering
b) Biopiracy
c) Biopatenting
d) Bioethics

Explanation: Unauthorized use of genetic material or bio-resources constitutes biopiracy. Biopatenting legally protects inventions, bioethics provides moral guidance, and bioengineering involves technology application. Option (b) is correct.

Q5. Single Correct Answer: Laws preventing commercialization without consent protect against:
a) Bioethics violations
b) Biopiracy
c) Bioengineering errors
d) Biopatenting conflicts

Explanation: Legal frameworks prevent biopiracy by stopping unauthorized commercialization of bio-resources or traditional knowledge. Bioethics guides ethical research, bioengineering applies technology, and biopatenting protects inventions. Option (b) is correct.

Q6. Single Correct Answer: Indigenous knowledge about plant properties is safeguarded to avoid:
a) Biopatenting
b) Biopiracy
c) Bioengineering
d) Bioethics

Explanation: Indigenous knowledge is vulnerable to exploitation; laws prevent biopiracy ensuring fair benefit sharing. Biopatenting protects inventions, bioengineering applies technology, and bioethics governs ethical conduct. Option (b) is correct.

Q7. Assertion-Reason:
Assertion (A): Biopiracy is unethical exploitation of bio-resources.
Reason (R): Intellectual property rights and laws prevent unauthorized use.

a) Both A and R are true, R explains A
b) Both A and R are true, R does not explain A
c) A is true, R is false
d) A is false, R is true

Explanation: Both assertion and reason are correct. Biopiracy involves unethical exploitation, and intellectual property laws protect bio-resources against unauthorized use, ensuring ethical and legal compliance. Option (a) is correct.

Q8. Matching Type: Match term with definition:
Column - I: a) Biopiracy, b) Biopatenting, c) Bioethics, d) Bioengineering
Column - II: i) Engineering application to biology, ii) Unauthorized exploitation, iii) Ethical study, iv) Legal protection of inventions

Options:
1. a-ii, b-iv, c-iii, d-i
2. a-iv, b-iii, c-ii, d-i
3. a-iii, b-ii, c-i, d-iv
4. a-i, b-ii, c-iv, d-iii

Explanation: Correct match: a-ii (Biopiracy: unauthorized exploitation), b-iv (Biopatenting: legal protection), c-iii (Bioethics: ethical study), d-i (Bioengineering: engineering application). Option (1) is correct.

Q9. Fill in the Blanks: The unauthorized use of biological resources without consent is called ________.
a) Biopatenting
b) Bioengineering
c) Biopiracy
d) Bioethics

Explanation: Unauthorized exploitation of bio-resources or traditional knowledge is termed biopiracy. Biopatenting legally protects inventions, bioengineering applies technology, and bioethics governs moral conduct. Option (c) is correct.

Q10. Choose the correct statements:
1) Biopiracy involves illegal exploitation of bio-resources
2) Biopatenting grants legal protection for inventions
3) Bioethics ensures ethical compliance in research
4) Bioengineering prevents unauthorized use

a) 1, 2, 3
b) 1, 3, 4
c) 2, 3, 4
d) 1, 2, 4