Differential plasmid rescue from transgenic mouse DNAs into Escherichia coli methylation-restriction mutants.
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Plasmids comprising transgene insertions in four lines of transgenic mice have been retrieved by plasmid rescue into a set of Escherichia coli strains with mutations in different members of the methylation-dependent restriction system (MDRS). Statistical analysis of plasmid rescue frequencies has revealed that the MDRS loci detect differential modifications of the transgene insertions among mouse lines that show distinctive patterns of transgene expression. Plasmids in mice that express hybrid insulin transgenes during development can be readily cloned into E. coli strains carrying mutations in two of the MDRS loci, mcrA and mcrB. In mice in which transgene expression is inappropriately delayed into adulthood, plasmids can only be cloned into E. coli that carry mutations in all known MDRS activities. Differential cloning frequencies in the presence or absence of the various methylation-dependent restriction genes represent a further way to distinguish regions of mammalian chromosomes. These multiply deficient E. coli strains will also facilitate the molecular cloning of modified chromosomal DNA. IMAGES:
