Multicopy plasmid modification with phage lambda Red recombineering.
Recombineering, in vivo genetic engineering using the bacteriophage lambda Red generalized recombination system, was used to create various modifications of a multicopy plasmid derived from pBR322. All genetic modifications possible on the Escherichia coli chromosome and on bacterial artificial chromosomes (BACs) are also possible on multicopy plasmids and are obtained with similar frequencies to their chromosomal counterparts, including creation of point mutations (5-10% unselected frequency), deletions and substitutions. Parental and recombinant plasmids are nearly always present as a mixture following recombination, and circular multimeric plasmid molecules are often generated during the recombineering.
Thomason
Lynn C
LC
Gene Regulation and Chromosome Biology Laboratory, Building 539, Room 243, National Cancer Institute at Frederick, Frederick, MD 21702, USA. lthomason@ncifcrf.gov
Costantino
Nina
N
Shaw
Dana V
DV
Court
Donald L
DL
D010582:Bacteriophage lambda / Q000235:genetics*; D022202:Chromosomes, Artificial, Bacterial; D002876:Chromosomes, Bacterial; D004926:Escherichia coli / Q000235:genetics; D017353:Gene Deletion; D005798:Genes, Bacterial; D005818:Genetic Engineering; D007612:Kanamycin / Q000494:pharmacology; D010957:Plasmids / Q000235:genetics*; D017354:Point Mutation; D011995:Recombination, Genetic
D007612:Kanamycin