FIG. 1.

Gene targeting of envoplakin locus. (a) Targeting strategy used to generate the envoplakin null allele. Top, 5′ end of the envoplakin gene. Black boxes represent the exons. Middle, the targeting vector inserts into the first exon of the envoplakin gene a cassette with stop codons in all reading frames, an internal ribosome entry site-dependent lacZ reporter gene, and a neomycin resistance gene. Bottom, diagnostic 5′ and 3′ end fragments to recognize correctly integrated targeting vector. (b) Genotyping the offspring of heterozygous crosses by Southern blotting. The −/− animals are homozygous for the targeted allele, as revealed by both the 5′ and 3′ probes. (c) RT-PCR analysis of mRNA from the skin of newborn mice. Both 5′-end primers (five first lanes from the left) that flank the deleted part of the gene and 3′-end primers (three right-hand lanes) from the last exon of the gene were used to verify the absence of mRNA in null animals. Primers for γ-actin were used as a control for the quality of the template RNA. Genotypes were determined by Southern blotting using the 5′-end probe.