Accumulated background variation among H2 mutant congenic strains: elimination through PCR-based genotyping of F-2 segregants

Many commercially and privately available congenic strains of laboratory an imals were founded decades ago and are likely to differ from one another by dozens of fixed mutational differences at background loci. This problem is often ignored despite growing evidence that such background variation exis ts. Eliminating this confounding variation can be largely accomplished by c rossing congenic strains to produce F-2 segregants that are homozygous (or heterozygous) for relevant genes. Discriminating F-2 homozygotes can be dif ficult when strain differences are minor, as are mutant mouse strains diffe ring at single major histocompatibility loci (H2 mutant congenics). Here, w e describe a two-step polymerase chain reaction (PCR) method utilizing hete roduplex analysis and sequence specific primers (SSP-PCR) that efficiently discriminates the F-2 progeny of two such H2 mutant congenic mice crosses ( bm1 X B6 and bm1 X bm3). A third H2 mutant cross cannot be resolved by hete roduplexing, but is discriminated (albeit less efficiently) with SSP-PCR al one. This sensitive application can be extended to any congenic mutant stra ins. (C) 2001 Elsevier Science B.V. All rights reserved.