Validations of floxed alleles by long-range genotyping PCR. (A) Illustration of the potential mosaicism of F0 founders. Although founders with 5′- and 3′-loxP insertions can be separately identified by genotyping PCR using 5′F + 5′R and 3′F + 3′R primers, this cannot distinguish whether loxP insertions are in cis or in trans. (B) Schematic to show the design and major steps of genotyping strategy. (C) DNA electrophoresis results of long-range PCR. The DNA bands enclosed in the green box were purified and cloned into vectors. (D–G) Representative genotyping results of bacterial colonies from cloning of the purified long-range PCR products for founder #5 of the Gm44386 gene (D), founder #7 for the Ak040954 gene (E), and founder #8 (F) and #11 (G) for the Plagl1 gene. (H) Summary of bacterial colony genotyping results.
