Generation of IGSF4-deficient mice and characterization of phenotype. (A) The trap vector, pU-21W, was inserted into the first intron of IGSF4. P1-2 (a primer set for the trap vector insertion point in the first intron; red arrows) and T1-2 (a primer set for the trap vector–specific sequence; orange arrows) were used for genotyping, and E1-2 (a primer set for the exon 1– and exon 2–specific sequence; blue arrows) was used for RT-PCR to detect endogenous IGSF4 mRNA. (B) PCR genotyping. Genomic DNA was extracted from mouse ear tissue. The WT (+/+) and trap (GT) alleles were detected with P1-2, but the homozygote (GT/GT) allele was not detected with this primer set (top). Total RNA was also extracted from mouse ear tissue, and the levels of IGSF4 mRNA were determined by using E1-2 (bottom). (C) Phenotypic comparison of IGSF4^+/+ and IGSF4^GT/GT mice. 4-wk-old and E18.5 WT and knockout littermates of size differences are shown. The organs from the IGSF4^GT/GT mice are slightly smaller than those of the WT littermates (He, heart; Ki, kidney; Sp, spleen; Th, thymus). Body weight was measured weekly. The data represent the mean ± SD (n = 8).