Targeted Disruption of the FucM Gene. (A) Schematic diagram for the targeted disruption of the FucM gene. The structures of the targeting vector, wild-type, and disrupted FucM alleles are shown. The solid boxes represent the exons (a total of six in the wild-type). The restriction sites used for constructing the targeting vector and for analysis by Southern blot are EcoRV, BssHI, ScaI, and KpnI. A puromycin expressing cassette was inserted between the EcoRV and ScaI sites, eliminating exons 2-4 and the EcoRV site of the WT gene. (B) Southern analysis for genomic DNA from the wild-type (+/+), heterozygous (+/-) and FucM null (-/-) mice. The expected sizes of the wild-type and disrupted FucM alleles are 10 and 16 Kb, respectively, which were detected by Southern blotting with EcoRV digestion and hybridization using the probe 1 shown in (A). (C) The wild-type (primer 1 and 2) and disrupted alleles (primer 3 and 4) were also detected by PCR.