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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.
Solution :
1. Theoretical Foundation of Restriction Endonucleases
Restriction endonucleases, often referred to as "molecular scissors," are enzymes capable of cleaving DNA molecules at specific recognition sequences known as palindromic sequences. The enzyme EcoRI, isolated from Escherichia coli RY13, is the canonical example. It recognizes the hexanucleotide sequence $5'-GAATTC-3'$ and performs a staggered cut between the $G$ and $A$ residues, generating "sticky ends" or cohesive termini.
2. Comparative Analysis of the Enzymatic Process
| Component | Description |
|---|---|
| Enzyme | EcoRI (Restriction Endonuclease) |
| Substrate | Double-stranded DNA (dsDNA) |
| Recognition Site | $5'-GAATTC-3'$ / $3'-CTTAAG-5'$ |
| Resultant Product | DNA fragments with single-stranded overhanging ends (sticky ends) |
3. Diagrammatic Representation of Restriction Cleavage
4. Sequential Mechanism of Action
- Step 1: Recognition [Binding]: The EcoRI enzyme scans the dsDNA molecule and binds specifically to the palindromic sequence $GAATTC$ [Per the specificity of DNA-protein interaction].
- Step 2: Cleavage [Hydrolysis]: The enzyme catalyzes the hydrolysis of the phosphodiester backbone between the $G$ and $A$ nucleotides on both strands [Mechanism: Nucleophilic attack on the phosphate group].
- Step 3: Dissociation: The DNA molecule is cleaved into two fragments. Because the cut is staggered, each fragment possesses an exposed single-stranded tail.
- Step 4: Product Formation: These ends are complementary to one another, allowing for hydrogen bonding with other DNA fragments cut by the same enzyme [Principle: Complementary Base Pairing].
Final Solution: The restriction enzyme EcoRI acts upon the substrate sequence $5'-GAATTC-3'$, cutting between $G$ and $A$ to produce DNA fragments featuring $5'$ overhangs, which facilitate the ligation process in recombinant DNA technology.
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.
- 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(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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