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. 2006 Dec 13;26(50):13089–13101. doi: 10.1523/JNEUROSCI.2855-06.2006

Figure 1.

Figure 1.

Generation and survival of Mint 1, 2, and 3 KO mice. A–C, Homologous recombination strategy for Mint 1, 2, and 3, respectively. The targeted exon (red box) was flanked by loxP sites (yellow arrowheads) to allow conditional removal of exon, a NEO for positive selection surrounded by FRT sites (green circles), and a diphtheria toxin (DT) was attached for negative selection. D–F, Immunoblot analysis of brain proteins from wild-type (+/+) and homozygous (−/−) mice. A–D, Homozygous floxed Mint 1 mutants (MT1) showed normal Mint 1 protein expression in the presence of NEO resistance cassette but were abolished when we crossed these mice with transgenic mice that express cre recombinase in the male germline (M1R). B, C, E, F, For homozygous floxed Mint 2 and 3 mutants (MT2 and MT3, respectively), the presence of the NEO resistance cassette abolished protein expression before floxed exon was excised. However, when NEO was excised by crossing with mice expressing FRT recombinase in the germline, both Mints 2 and 3 expressions returned as expected (M2F, M3F, respectively). In addition, Mints 2 and 3 protein expression were abolished when exon was removed by crossing with mice that express cre recombinase (M2R, M3R, respectively). In addition, we have created Mint KO lines in which protein expression was abolished with excision of both the targeted exon and NEO cassette (M1K, M2K, and M3K). G, Genotype analysis of offspring obtained from single Mint 1, 2, or 3 heterozygous matings show normal Mendelian ratio of genotypes. Genotype analysis of offspring obtained from Mint 1−/−/3+/− or Mint 2+/−/3−/− matings revealed normal Mendelian ratio of genotypes. However, interbreeding of Mint 1−/−/2+/− animals resulted in a Mendelian ratio of 5% instead of the expected 25% in double Mint 1/2 offsprings. Triple KO of all three Mints showed a Mendelian ratio similar to Mint 1/2 double KO mice. Sp, Spe; P, probe; M, Mfe1; BB, BstB1; B, BamH1; S, Sal1; EV, EcoRV; X, Xho1; Ns, Nsi1; N, Not1.