FIG. 1.

Generation and molecular analysis of Ets1^ΔVII mice. (A) Targeting strategy. The top row shows Ets1 genomic structure and restriction map of an Ets1 genomic clone isolated from a 129/Sv mouse genomic library. Restriction sites: A, ApaI; B, BamHI; C, ClaI; N, NotI; and X, XhoI. Primer binding sites are indicated by numbered arrows (p1, p2, p3). Three-prime flanking probe and 10.5-kb wild-type BamHI fragment are indicated. The second row from the top shows the structure of the Ets1^ΔVII targeting vector. A loxP element (black triangle) was cloned into a unique ApaI site 5′ to exon VII, and a pGH-1 G418 resistance cassette (Neo^r) and loxP element were cloned into a unique ClaI site 3′ to exon VII. The third row from the top shows the structure of the Cre conditional Ets1^ΔVII-targeted allele. The reduction in size of the 3′ BamHI fragment is indicated. The bottom row depicts a Cre recombined allele lacking exon VII. Genotyping primers are indicated. (B) Confirmation of gene targeting in ES cells by Southern blotting using a 3′ BamHI-XhoI probe to show 10.5-kb wild-type and 6.7-kb targeted alleles. (C) PCR verification of Cre recombination. Recombinant (R), targeted (T), and wild-type (W) alleles are indicated with primer pairs. Heterozygous targeted (Ta) and recombined (Re) cells and the targeting vector (TV) control are shown. (D) Western blot confirmation of protein expression from the targeted allele in isolated thymocytes and splenocytes. Twofold-greater total spleen protein versus total thymus protein was loaded due to reduced Ets1 expression in the spleen. Ets1 genotypes are given above each blot.