| 摘要 |
This invention pertains to a method of site-specific mutagenesis of DNA. The method can be used to mutagenize DNA, especially circular DNA (for example, virtually any plasmid), requiring only that the DNA carry a nonessential, unique recognition site for a restriction enzyme (restriction site). According to the method, a parental DNA to be mutated, containing a nonessential, unique restriction site, is used to generate progeny DNA containing the desired mutation but lacking the restriction site. The non-mutant parental-type DNA and the mutant progeny DNA are treated with any enzyme that cleaves at the restriction site to cleave the parental-type DNA, leaving the mutant DNA uncleaved. Mutant progeny DNA is selected by transforming cells with the enzyme-treated DNA under conditions such that the cells are transformed with uncleaved DNA at a higher efficiency than cleaved DNA with the result that the majority of transformants carry the mutant DNA. In preferred embodiments, the DNA is circular. The restriction enzyme treatment linearizes parental DNA which retains the nonessential, unique restriction site, leaving mutant DNA which lacks the site circular. Circular DNA transforms host cells more efficiently than linear DNA.
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