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. 2000 Dec 1;106(11):1361–1371. doi: 10.1172/JCI9154

Figure 4.

Figure 4

PCR analysis of genomic DNA prepared from tissues of V2R mutant mice to verify Cre-mediated excision of the neo gene. Homozygous male EIIa-cre mice were bred with female V2R+/– mutant mice. Resulting V2R+/– female offspring were then interbred with wild-type male littermates to achieve germline transmission of the neo deletion. To verify the ablation of the neo gene, genomic DNA was prepared from tail (Ta), kidney (Ki), and liver (Li) samples derived from the resulting pups and subjected to PCR genotyping. The two PCR primers used for this analysis were chosen to flank the loxP-enclosed neo cassette and were located at the exon 2/intron 2 (5′-gaagctcctctggaaagtggt-3′) and intron 2/exon 3 (5′-tcctatgaagaagagagaccag-3′) junctions (see Figure 1a), respectively. After Cre-mediated excision of the neo gene (note that one 34-bp loxP sequence remains in the genome after the excision of the neo cassette), the two primers amplify a 210-bp PCR fragment, whereas the corresponding wild-type fragment is 34 bp shorter (176 bp). The absence of neo sequences in all samples was confirmed by the use of internal neo primers (data not shown). The DNAs analyzed here were taken from three different mouse pups of the indicated V2R genotypes.