title: A new site-specific recombinase-mediated system for targeted multiple genomic deletions employing chimeric loxP and mrpS sites.
abstract: A newly designed site-specific recombination system is presented which allows multiple targeted markerless deletions. The most frequently used tool for removing selection markers or to introduce genes by recombination-mediated cassette exchange is the Cre/loxP system. Many mutant loxP sites have been created for this purpose. However, this study presents a chimeric mutant loxP site denoted mroxP-site. The mroxP site consists of one Cre (loxP/2) and one MrpA (mrpS/2) binding site separated by a palindromic 6-bp spacer sequence. Two mroxP-sites can be recombined by Cre recombinase in head-to-tail as well as in head-to-head orientation. In the head-to-head orientation and the loxP half-sites inside, Cre removes the loxP half-sites during site-specific recombination, creating a new site, mrmrP. The new site is essentially a mrpS site with a palindromic spacer and cannot be used by Cre for recombination anymore. It does, however, present a substrate for the recombinase MrpA. This new system has been successfully applied introducing multiple targeted gene deletions into the Escherichia coli genome. Similar to Cre/loxP and FLP/FRT, this system may be adapted for genetic engineering of other pro- and eukaryotes.
authors.affiliation: Institut für Industrielle Genetik, Allmandring 31, Universität Stuttgart, 70569, Stuttgart, Germany.
authors.forename: Lydia
authors.initials: L
authors.lastname: Warth
authors.forename: Josef
authors.initials: J
authors.lastname: Altenbuchner
mesh_terms: D001483:Base Sequence; D002876:Chromosomes, Bacterial; D004926:Escherichia coli / Q000235:genetics; D017353:Gene Deletion; D023281:Genomics; D019426:Integrases / Q000378:metabolism*; D010957:Plasmids; D016133:Polymerase Chain Reaction; D011995:Recombination, Genetic; D012689:Sequence Homology, Nucleic Acid
chemical_list: C045073:Cre recombinase; D019426:Integrases