Figure 6. Cre-mediated recombination of the Smad4^f allele in spermatogenic cells of double transgenic Kera-Cre/Smad4^f/wt mouse. (A) Histograms of flow cytometry of testicular cell suspension obtained from a 60-day-old double transgenic Kera-Cre/Smad4^f/w mouse showed a distribution of 65%, 9%, and 17% for 1N, 2N, and 4N cells, respectively. PCR of genomic DNA from 1N and 4N cells showed excision of the Smad4^f allele but not from 2N cells. (B) Histogram showed enrichment of 2N cells by Percoll gradients as described in Methods. PCR analysis of these enriched 2N cells demonstrated excision of the Smad4^f allele. Primers b and c identify wild type Smad4 (390 bp) and floxed Smad4^f allele (450 bp) whereas primers a and c identify the excised Smad4^f allele (500 bp). TC, testicular cells; M, DNA marker.