Isolation of Cosmid and BAC DNA from E. coli
Cosmid and bacterial artificial chromosome (BAC) systems have been developed for the cloning of large DNA inserts averaging 40 kb and 130 kb (range: 90–300 kb), respectively. The resulting clones are more stable than yeast artificial chromosomes (YACs) and rarely chimeric, which makes them excellent tools for the generation of contiguous physical maps. The use of such large-insert clones considerably increases the rate of complex genome mapping and sequencing. Numerous standard protocols have been developed to isolate supercoiled plasmids, but of these, the alkaline lysis protocol remains the most suitable approach to isolate large-insert clones such as cosmids and BACs.
KeywordsVortex Glycerol Phenol EDTA Chloroform