Plasmid Isolation Protocol @IITianTheory2Tech

Plasmid isolation is based on the alkaline lysis method, which selectively separates plasmid DNA from chromosomal DNA. Bacterial cells are first harvested and resuspended in a buffer containing Tris, EDTA, and glucose to maintain stability. Lysis is achieved using SDS and NaOH, which disrupt membranes and denature DNA and proteins. Upon neutralization with potassium acetate, chromosomal DNA and proteins precipitate, while plasmid DNA remains in solution. After centrifugation, plasmid DNA is recovered from the supernatant, precipitated using alcohol, washed with ethanol, and finally dissolved in TE buffer. This method is cost-effective and widely used in molecular biology laboratories.

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