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Q11(a):
Explain briefly (a) PCR
Solution :
Theoretical Foundation: Polymerase Chain Reaction (PCR)
PCR is an *in vitro* method for enzymatic amplification of specific DNA sequences. Developed by Kary Mullis, it utilizes the thermostable DNA polymerase, Taq polymerase, derived from the thermophilic bacterium Thermus aquaticus, to exponentially increase the copy number of a target DNA fragment. [Principle: Molecular replication via thermal cycling].
Step 1: Denaturation
The double-stranded DNA (dsDNA) template is subjected to high temperatures (approximately $94^{\circ}\text{C} - 96^{\circ}\text{C}$) for 30–60 seconds. This thermal energy disrupts the hydrogen bonds between complementary nitrogenous bases. [By the principle of thermal denaturation of nucleic acids].
Result: Conversion of dsDNA into two single-stranded DNA (ssDNA) templates.
Step 2: Annealing
The reaction mixture is cooled to $50^{\circ}\text{C} - 65^{\circ}\text{C}$ for 20–40 seconds. Two short, synthetic oligonucleotide primers (typically 15–30 nucleotides long) hybridize to their complementary sequences flanking the target region on the ssDNA templates. [By the principle of complementary base pairing].
Step 3: Extension (Polymerization)
The temperature is raised to $72^{\circ}\text{C}$ (the optimal temperature for Taq polymerase activity). The polymerase enzyme adds deoxyribonucleoside triphosphates (dNTPs) to the $3'$-OH end of the primers, synthesizing a new DNA strand in the $5' \rightarrow 3'$ direction. [Per the mechanism of DNA-dependent DNA polymerase].
Schematic Representation of PCR Cycles
Amplification Efficiency
PCR follows exponential kinetics. If $n$ represents the number of cycles, the total number of DNA molecules produced is defined by the formula $2^n$.
| Cycle Number | DNA Molecules |
|---|---|
| 0 | 1 |
| 1 | 2 |
| 2 | 4 |
| 10 | 1,024 |
| 30 | ~1,000,000,000 |
Final Solution: Polymerase Chain Reaction (PCR) is a tripartite thermal process consisting of denaturation, annealing, and extension, which uses Taq polymerase to achieve the exponential amplification of a specific DNA segment, theoretically yielding $2^n$ copies after $n$ cycles.
More Questions from Class 12 Biology Biotechnology: Principles and Processes EXERCISES
- Q1: Can you list 10 recombinant proteins which are used in medical practice? Find out where they are used as therapeutics (use the internet).
- Q1: Can you list 10 recombinant proteins which are used in medical practice? Find out where they are used as therapeutics (use the internet).
- Q2: Make a chart (with diagrammatic representation) showing a restriction enzyme, the substrate DNA on which it acts, the site at which it cuts DNA and the product it produces.
- Q2: Make a chart (with diagrammatic representation) showing a restriction enzyme, the substrate DNA on which it acts, the site at which it cuts DNA and the product it produces.
- Q3: From what you have learnt, can you tell whether enzymes are bigger or DNA is bigger in molecular size? How did you know?
- Q3: From what you have learnt, can you tell whether enzymes are bigger or DNA is bigger in molecular size? How did you know?
- Q4: What would be the molar concentration of human DNA in a human cell? Consult your teacher.
- Q4: What would be the molar concentration of human DNA in a human cell? Consult your teacher.
- Q5: Do eukaryotic cells have restriction endonucleases? Justify your answer.
- Q5: Do eukaryotic cells have restriction endonucleases? Justify your answer.
- Q6: Besides better aeration and mixing properties, what other advantages do stirred tank bioreactors have over shake flasks?
- Q6: Besides better aeration and mixing properties, what other advantages do stirred tank bioreactors have over shake flasks?
- Q7: Collect 5 examples of palindromic DNA sequences by consulting your teacher. Better try to create a palindromic sequence by following base-pair rules.
- Q7: Collect 5 examples of palindromic DNA sequences by consulting your teacher. Better try to create a palindromic sequence by following base-pair rules.
- Q8: Can you recall meiosis and indicate at what stage a recombinant DNA is made?
- Q8: Can you recall meiosis and indicate at what stage a recombinant DNA is made?
- Q9: Can you think and answer how a reporter enzyme can be used to monitor transformation of host cells by foreign DNA in addition to a selectable marker?
- Q9: Can you think and answer how a reporter enzyme can be used to monitor transformation of host cells by foreign DNA in addition to a selectable marker?
- Q10(a): Describe briefly the following: (a) Origin of replication
- Q10(a): Describe briefly the following: (a) Origin of replication
- Q10(b): Describe briefly the following: (b) Bioreactors
- Q10(b): Describe briefly the following: (b) Bioreactors
- Q10(c): Describe briefly the following: (c) Downstream processing
- Q10(c): Describe briefly the following: (c) Downstream processing
- Q11(a): Explain briefly (a) PCR
- Q11(b): Explain briefly (b) Restriction enzymes and DNA
- Q11(b): Explain briefly (b) Restriction enzymes and DNA
- Q11(c): Explain briefly (c) Chitinase
- Q11(c): Explain briefly (c) Chitinase
- Q12(a): Discuss with your teacher and find out how to distinguish between (a) Plasmid DNA and Chromosomal DNA
- Q12(a): Discuss with your teacher and find out how to distinguish between (a) Plasmid DNA and Chromosomal DNA
- Q12(b): Discuss with your teacher and find out how to distinguish between (b) RNA and DNA
- Q12(b): Discuss with your teacher and find out how to distinguish between (b) RNA and DNA
- Q12(c): Discuss with your teacher and find out how to distinguish between (c) Exonuclease and Endonuclease
- Q12(c): Discuss with your teacher and find out how to distinguish between (c) Exonuclease and Endonuclease
CBSE Solutions for Class 12 Biology Biotechnology: Principles and Processes
Chapters in CBSE - Class 12 Biology
- Reproduction in Organisms
- Human Reproduction
- Reproductive Health
- Principles of Inheritance and Variation
- Molecular Basis of Inheritance
- Evolution
- Human Health and Disease
- Strategies for Enhancement in Food Production
- Microbes in Human Welfare
- Biotechnology and Its Applications
- Organisms and Populations
- Ecosystem
- Biodiversity and Conservation
- Environmental Issues
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