RRC ID 51564
Author Lee G, Saito I.
Title Role of nucleotide sequences of loxP spacer region in Cre-mediated recombination.
Journal Gene
Abstract The Cre recombinase mediates precise site-specific recombination between a pair of loxP sequences through an intermediate containing Holiday junction. The recombination junction in the loxP sequence is located within the asymmetric 8-nucleotide spacer region. To examine the role of each nucleotide sequence of the spacer region in the recombination process, we synthesized a complete set of 24 loxP spacer mutants with single-base substitutions and 30 loxP spacer mutants with double-base substitutions. Each synthesized loxP mutant was ligated at both ends of a linear DNA or to one end of a DNA-containing wild-type loxP at the other end and their recombination efficiencies were analyzed with an in vitro system. The sequence identity of the right two nucleotides and left four nucleotides in the central six bases of the spacer region was found to be essential for formation and resolution, respectively, of the intermediate product. Furthermore, even when homology was maintained, the recombination efficiencies were lower than that of wild-type loxP and varied among mutants. Based on this knowledge, we identified two loxP mutants with double-base substitutions, mutants 5171 and 2272, which recombine efficiently with an identical mutant but not with the other mutant or wild-type loxP.
Volume 216(1)
Pages 55-65
Published 1998-8-17
DOI 10.1016/s0378-1119(98)00325-4
PII S0378-1119(98)00325-4
PMID 9714735
MeSH Base Sequence / physiology Binding Sites / genetics Cell Line, Transformed DNA / chemistry DNA / genetics DNA / physiology* Gene Expression / genetics Humans Integrases / genetics Integrases / metabolism* Molecular Sequence Data Mutation / genetics Point Mutation / genetics Recombination, Genetic Substrate Specificity Viral Proteins*
IF 2.638
Times Cited 227
Resource
DNA material pUVwV (RDB04396) pUSwS (RDB04397) pULwV (RDB04398).