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Fig. 1. Targeted disruption of the Tbx20 locus. (A) Schematic representation of the targeting strategy. Restriction map of the wild-type locus with boxes representing the first four exons of Tbx20; coding regions are shown in black, non-coding in white. Fragments used as RFLP probes are shown. The KpnI-EcoRV fragment designated as 5' detects the KpnI-RFLP shown in B. The HincII-RFLP shown in C is detected by the BamHI-KpnI fragment labelled as 3'. Only HincII sites relevant for RFLP analysis are shown. B, BamHI; E, EcoRI; H, HincII; K, KpnI; N, NcoI; RV, EcoRV; neo, loxP-flanked neomycin selection cassette. (B) Southern blot analysis of KpnI-digested genomic DNA extracted from E9.5 embryos derived from intercrosses of mice heterozygous for the mutant Tbx20 allele. Genotypes are indicated above each lane. The 4.8 kb and 8.2 kb bands represent the wild-type and the mutant allele, respectively. (C) Southern blot analysis of HincII-digested genomic DNA extracted from the same E9.5 embryos. Genotypes are indicated above each lane. The 10 kb and 8 kb bands represent the wild-type and the mutant allele, respectively. (D) In situ hybridization analysis of Tbx20 expression in wild-type (+/+) and Tbx20^–/– (–/–) embryos at E8.5 using an antisense riboprobe against the T-box shows complete absence of Tbx20 mRNA in the mutant embryo.

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