Abstract
This chapter explains the triton–lysozyme method of plasmid DNA preparation, which is used to prepare plasmid DNA from E. coli . The bacteria are partially lysed with lysozyme and triton, allowing the plasmid to escape the cell wall into the supernatant. The larger E. coli chromosomal DNA is trapped in the cell wall. The lysate is cleared of cell debris, and the plasmid-containing supernatant is banded in a CsCl/ ethidium bromide equilibrium gradient to purify closed circular DNA from any sheared linear plasmid or E. coli DNA fragments. The chapter discusses a commonly used method that generates highly purified plasmid DNA, free of RNA contamination, but requires a considerable amount of time and specialized ultracentrifuge equipment. The chapter lists the reagents that are required to carry out the procedure for the triton–lysozyme method of plasmid DNA preparation.
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