A genetic screen identifies novel non-compatible loxP sites.
The ability of the Cre/lox system to make precise genomic modifications is a tremendous accomplishment. However, recombination between cis-linked heterospecific lox sites limits the use of Cre- mediated exchange of DNA to systems where genetic selection can be applied. To circumvent this problem we carried out a genetic screen designed to identify novel mutant spacer-containing lox sites displaying enhanced incompatibility with the canonical loxP site. One of the mutant sites recovered appears to be completely stable in HEK293 cells constitutively expressing Cre recombinase and supports recombinase-mediated cassette exchange (RMCE) in bacteria and mammalian cell culture. By preventing undesirable recombination, these novel lox sites could improve the efficiency of in vivo gene transfer.
Department of Molecular, Cellular and Developmental Biology, CB347, University of Colorado, Boulder, CO 80309-0347, USA.
Stephen J
SJ
Langer
A Paiman
AP
Ghafoori
Marshall
M
Byrd
Leslie
L
Leinwand
D001419:Bacteria / Q000235:genetics; D001665:Binding Sites / Q000235:genetics; D002460:Cell Line; D004247:DNA / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D005838:Genotype; D006801:Humans; D019426:Integrases / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D009841:Oligonucleotides / Q000235:genetics; D010957:Plasmids / Q000235:genetics; D011995:Recombination, Genetic / Q000235:genetics*
D009841:Oligonucleotides; D004247:DNA; D019426:Integrases