How can we differ plasmid dna and genomic dna from total cell dna?
Most methods for isolation of DNA rely on disruption of cells in the presence of strong denaturants. Disruption may be by freezing and fracturing cells by grinding or blending or by chemical lysis with strong alkali. The denaturants are essential to inactivate exogeneous and endogenous nucleases which would otherwise degrade the DNA. Examine the components of several DNA extraction buffers and decide the purpose of each chemical. Plasmid DNA Isolation by Alkaline Lysis Plasmids are non-obligate, circular, extrachromosomal bacterial replicons. Plasmid DNA isolation requires separation of this DNA from the chromosomal DNA in the bacterial cell as well as from the polysaccharides, lipids and proteins that constitute the cell. Subsequent manipulation, especially enzymatic modification, of the plasmid DNA requires that it be free of these impurities. In the protocols below, cells are lysed by strong alkli (NaOH) and proteins are denatured by strong alkali and strong detergent (SDS). The det
