Gene doctoring: a method for recombineering in laboratory and pathogenic Escherichia coli strains.

ABSTRACT:

Background

Homologous recombination mediated by the lambda-Red genes is a common method for making chromosomal modifications in Escherichia coli. Several protocols have been developed that differ in the mechanisms by which DNA, carrying regions homologous to the chromosome, are delivered into the cell. A common technique is to electroporate linear DNA fragments into cells. Alternatively, DNA fragments are generated in vivo by digestion of a donor plasmid with a nuclease that does not cleave the host genome. In both cases the lambda-Red gene products recombine homologous regions carried on the linear DNA fragments with the chromosome. We have successfully used both techniques to generate chromosomal mutations in E. coli K-12 strains. However, we have had limited success with these l

SUBMITTER: Lee DJ 

PROVIDER: S-EPMC2796669 | biostudies-literature | 2009 Dec

REPOSITORIES: biostudies-literature