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Q11(b):
Explain briefly (b) Restriction enzymes and DNA
Solution :
Definition and Functional Role of Restriction Enzymes
Restriction enzymes, scientifically categorized as restriction endonucleases, are proteins that function as molecular "scissors." They identify specific nucleotide sequences within a DNA molecule, known as recognition sequences or restriction sites, and catalyze the hydrolysis of phosphodiester bonds at these precise locations [Per the biochemical mechanism of phosphodiester bond cleavage].
Step 1: Mechanism of Action
The process of DNA cleavage occurs via the following stages:
- Recognition: The enzyme scans the DNA duplex and binds to a specific palindromic sequence (a sequence that reads the same 5' to 3' on both strands).
- Binding: The enzyme interacts with the DNA backbone using hydrogen bonding and electrostatic interactions.
- Cleavage: The enzyme catalyzes the nucleophilic attack of a water molecule on the phosphate group of the DNA backbone, severing the strand.
Step 2: Types of Cuts (Blunt vs. Sticky Ends)
Restriction enzymes produce two primary types of fragments based on the location of the cut:
| Type of Cut | Mechanism | Biological Advantage |
|---|---|---|
| Sticky Ends | Staggered cuts within the recognition site, leaving short, single-stranded overhangs. | Allows for easy base-pairing with complementary sequences via DNA ligase [By Complementary Base Pairing]. |
| Blunt Ends | Cuts in the center of the recognition site, leaving no overhangs. | Often used in cloning where specific terminal ends are not required, though ligation is less efficient. |
Step 3: Relationship between Enzymes and DNA
In recombinant DNA technology, the interaction between restriction enzymes and DNA serves as the fundamental prerequisite for gene cloning:
- DNA Fragment Generation: Genomic DNA is treated with enzymes to produce fragments containing the target gene.
- Vector Preparation: The same restriction enzyme is used on a vector (e.g., plasmid) to generate identical, complementary "sticky" ends.
- Annealing: Because the enzyme created complementary sequences, the gene of interest and the vector DNA spontaneously pair through hydrogen bonding [Per Chargaff’s Rules].
- Ligation: The enzyme DNA Ligase completes the process by sealing the gaps, creating a recombinant DNA molecule.
Final Solution: Restriction enzymes are site-specific endonucleases that recognize unique palindromic DNA sequences and induce cleavage to create either sticky or blunt ends. This enzymatic action is the essential mechanism for isolating DNA fragments and integrating them into vectors to produce recombinant DNA constructs.
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(a): Explain briefly (a) PCR
- 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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