title: Transfer of Large Contiguous DNA Fragments onto a Low Copy Plasmid or into the Bacterial Chromosome.
abstract: Bacterial pathogenicity islands and other contiguous operons can be difficult to clone using conventional methods due to their large size. Here we describe a robust 3-step method to transfer large defined fragments of DNA from virulence plasmids or cosmids onto smaller autonomously replicating plasmids or directly into defined sites in the bacterial chromosome that incorporates endogenous yeast and λ Red homologous recombination systems. This methodology has been successfully used to isolate and integrate at least 31 kb of contiguous DNA and can be readily adapted for the recombineering ofE. coliand its close relatives.
authors.affiliation: Department of Medicine, Division of Infectious Diseases, Massachusetts General Hospital, Cambridge, USA; Department of Microbiology and Immunobiology, Harvard Medical School, Boston, USA.
authors.forename: Analise Z
authors.initials: AZ
authors.lastname: Reeves
authors.affiliation: Department of Medicine, Division of Infectious Diseases, Massachusetts General Hospital, Cambridge, USA; Department of Microbiology and Immunobiology, Harvard Medical School, Boston, USA.
authors.forename: Cammie F
authors.initials: CF
authors.lastname: Lesser