Figure 1.
Targeted disruption of chicken FANCC loci in DT40 cells. (A) Schematic representation of partial chicken FANCC locus, the gene disruption construct, and the configuration of targeted allele. S, SacI; B, BamHI; H, HindIII. Only a single exon was deleted by the gene disruption. Arrowheads indicate the positions of primers used in RT–PCR. (B) Southern blot analysis of SacI-digested genomic DNA from cells with indicated genotypes using flanking probe as shown in (A). WT, wild type. (C) RT–PCR analysis of total RNA from wild-type and fancc cells. Primers were designed from FANCC sequences of upstream and downstream exons as shown in (A). As a control, the entire coding region of chicken Rad51 was amplified from each RT product. (D) Western blot analysis of whole-cell lysate prepared from wild-type and fancc cells probed with anti-chicken FANCD2 serum. Cells were treated with MMC (500 ng/ml) for 1 h, washed, and lysed 6 h later. L or S, long or short form of FANCD2, respectively.