Figure 1.

Generation of Cld-5–deficient mice. (a) Restriction maps of the wild-type allele, the targeting vector, and the targeted allele of the mouse Cld-5 gene. Only one exon covers the whole open reading frame of Cld-5. The targeting vector contained the pgk neo cassette in its middle portion to delete the exon in the targeted allele. The positions of the 5′ and 3′ probes for Southern blotting are indicated as bars. E, EcoRI; Sa, SacI; K, KpnI; N, NcoI; and S, Sse8387I. (b) Genotype analyses by Southern blotting of Eco RI-digested genomic DNA from wild-type (+/+), heterozygous (+/−), and homozygous (−/−) mice for the mutant Cld-5 gene allele. Southern blotting with 5′ and 3′ probes yielded an 8.4-kb band from the wild-type allele, and a 4.7- and 3.7-kb band from the targeted allele, respectively. (c) Loss of Cld-5 mRNA in the brain of Cld-5 ^−/− mice examined by RT-PCR. As a control, the hypoxanthine phosphoribosyl transferase gene was equally amplified in all samples. (d) Newborn Cld-5 ^+/+ and Cld-5 ^−/− mice. Homozygous Cld-5–deficient mice were born in the expected Mendelian ratios, and looked normal macroscopically. However, their movements gradually ceased, and they all died within 10 h of birth.