Creation of a <i>Tbx3</i> conditional null allele.

<p>A) Schematics of wild type mouse <i>Tbx3</i> locus and gene targeted alleles. LoxP sites were inserted 3.3 kb 5′ of the translational start and in intron 1 along with an Frt-flanked neomycin resistance selection cassette to create the <i>Tbx3^floxneo</i> allele. FLPe was used to remove the neomycin resistance selection cassette from animals bearing the <i>Tbx3^floxneo</i> allele to generate the <i>Tbx3^flox</i> allele; Cre-mediated recombination generates the <i>Tbx3^Δflox</i> allele. B) Quantitative real-time PCR analysis of cDNA from reverse transcribed <i>Tbx3</i> mRNA in e10.5 mouse embryos. Bar graphs compare levels of transcripts detected using primer sets depicted in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0067841#pone-0067841-g001" target="_blank">Figure 1A</a>. In <i>Tbx3^Δflox/+</i> heterozygotes, 50% of wild type levels of exon 1/2 (gray bar) and 5/6 (black bar) containing transcripts are present as expected from the single wild type allele. In <i>Tbx3^{Δflox/Δflox}</i> homozygotes, no exon 1/2 or 5/6 containing transcripts are present (none). C,D) Confocal micrographs of sectioned E10.0 limb buds after fluorescent immunohistochemical detection of Tbx3 using an antibody to the C-terminus. C1-4) <i>Tbx3^+/+</i> limb bud. C1) Merged color view of DAPI and FITC channels at low magnification. C2-4) Close up of white boxed region in C1. C2) DAPI channel showing DNA immunoreactivity. C3) FITC channel showing Tbx3 immunoreactivity. C4) Merged color view. D1-4) <i>Tbx3^{Δflox/Δflox}</i>; <i>PrxCre</i> limb bud. D1) Merged color view of DAPI and FITC channels at low magnification. D2-4) Close up of white boxed region in C1. D2) DAPI channel showing DNA immunoreactivity. D3) FITC channel showing lack of Tbx3 immunoreactivity. D4) Merged color view.