Fig. 1. Targeted disruption of mouse Tbx3 to produce the Tbx3^tm1Pa allele by homologous recombination in embryonic stem (ES) cells, followed by in vitro Cre-mediated excision. (A) The Tbx3 cDNA and exons 1-6 of the endogenous locus are shown. The T-box is shaded, coding exons are shaded or open boxes, the 5' and 3' UTR are shown as black boxes, and loxP sites are black arrowheads. The targeting construct, containing 7.2 kb of total homology, replaced three coding exons of the T-box with a loxP-flanked hsv-tk PGK-neomycin resistance cassette. A ß-actin-diptheria toxin (ß-actin DT) cassette (Maxwell et al., 1987) was used for negative selection. (B) Southern analysis of BamHI-digested genomic DNA from wild-type and targeted ES cell clones using the 5' external (ext) and 3' internal (int) probes indicated in A to distinguish a 15 kb endogenous band (+) from a 6.1 or 9.3 kb targeted band (–^t) with the 5' or 3' probes, respectively. (C) PCR genotyping of mice using primers indicated in A to distinguish a 330 bp wild-type and a 516 bp mutant band from the Cre-excised allele, Tbx3^tm1Pa (–^e).

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