Generation of mice with disruption of the Mcfd2 allele. (A) Schematic diagram of the Mcfd2 allele–targeting strategy. Locations of restriction enzymes, Southern blot probe, and genotyping primers are indicated. (B) Southern blot analysis of a targeted ES cell clone. Expected sizes of restriction fragments are (WT and KO) NsiI, 12.7 and 19.2 kb; ScaI, 12.5 and 7.4 kb; AflII, 9 and 8.5 kb; and BglII, 8.7 and 7.1 kb. Arrows indicate DNA ladder bands (kb). (C) Three-primer PCR genotyping results. The higher band in the heterozygous sample represents heteroduplex DNA. (D) Western blot analysis of liver lysates of WT, Mcfd2^−/−, and Lman1^gt1/gt1/Mcfd2^−/− mice. Three individual mice from each genotype were analyzed. A, AflII; B, BglII; N, NsiI; S, ScaI.