17
causes DNA strands to separate, and this method is used to denature genomic DNA during plasmid
isolation.
EXPLANATION
Restriction Endonucleases, Modifying Enzymes
- Alkaline Lysis Principle (Birnboim & Doly, 1979):
- During plasmid extraction, bacteria are lysed using a solution of $0.2\text{ N NaOH}$ and $1\%\text{ SDS}$, creating an alkaline pH ($\approx 12.0 - 12.5$).
- Denaturation Behavior:
- Chromosomal DNA: High alkalinity breaks hydrogen bonds between base pairs, causing large, sheared, linear chromosomal DNA strands to completely separate and denature irreversibly.
- Plasmid DNA: Small, covalently closed circular (ccc) plasmids denature partially but remain topologically intertwined.
- Neutralization: Rapid neutralization with acidic potassium acetate ($pH \approx 4.8 - 5.2$) allows plasmid circles to renature correctly, whereas genomic DNA tangles irreversibly into an insoluble aggregate that precipitates with SDS-protein complexes.
- nalysis:
- (Alkalinity; irreversibly): Correct.
Correct option: (2) Alkalinity; irreversibly
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