A self-deleting Cre-lox-ermAM cassette, Cheshire, for marker-less gene deletion in Streptococcus pneumoniae.
Although targeted mutagenesis of Streptococcus pneumoniae is readily accomplished with the aid of natural genetic transformation and chimeric donor DNA constructs assembled in vitro, the drug resistance markers often employed for selection of recombinant products can themselves be undesirable by-products of the genetic manipulation. A new cassette carrying the erythromycin-resistance marker ermAM that can be used as a temporary marker for selection of desired recombinants is described. The cassette may subsequently be removed at will by virtue of an embedded fucose-regulated Cre recombinase gene and terminal lox66 and lox71 Cre recognition sites, with retention of 34bp from the cassette as an inert residual double-mutant lox72 site.
Laboratory for Molecular Biology, Department of Biological Sciences, University of Illinois at Chicago, Chicago, IL 60607, USA.
Liming
L
Weng
Indranil
I
Biswas
Donald A
DA
Morrison
D001483:Base Sequence; D003001:Cloning, Molecular / Q000379:methods; D005643:Fucose; D017353:Gene Deletion; D005819:Genetic Markers; D019426:Integrases; D008957:Models, Genetic; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional / Q000379:methods*; D011401:Promoter Regions, Genetic; D011995:Recombination, Genetic; D016415:Sequence Alignment; D013296:Streptococcus pneumoniae / Q000235:genetics*
D005819:Genetic Markers; D005643:Fucose; C045073:Cre recombinase; D019426:Integrases