Replacement vectors with genomlc DNA originating from different mouse strains were used to introduce site-specific mutations into the creatine kinase M (CKM) gene of mouse embryonic stem (ES) cells. Here we demonstrate that in mouse strain 129-derived ES cells, the gene is at least 25-fold more efficiently targeted with an Isogenic, 129-derived vector (129-pRV8.3) than with a nonisogenic, BALB/c-specific vector (BALB/c-pRV8.3). The two targeting constructs were identical except for allellc differences which were typed by partial sequencing. These included base pair mismatches (2%) and a polymorphic [GTC]-repeat length variation. Both in separate transfectlons as well as in cotransfections with mixed vectors, homologous disruption of the CKM gene resulted uniquely from the 129-isogenic DNA. Our data confirm earlier observations on requirements for homologous recombination in pro- and eukaryotic systems and indicate that targeting of the CKM locus is highly sensitive to small sequence differences between cognate segments in the endogenous and incoming DNA.
|Original language||English (US)|
|Number of pages||6|
|Journal||Nucleic acids research|
|State||Published - Aug 11 1992|
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