FIGURE 5.

TYW3 locus identification. (A) Schematic of the TYW3 transgene, showing positions of primers used for TLA. Both primer sets were designed within the Thy1 sequence. (B) Genome-wide TLA coverage using Thy1 primer set 1. Peak at Chromosome 9 shows endogenous Thy1 (black circle) and peak at Chromosome 1 shows inserted sequence (red circle). (C) Genome-wide TLA coverage using Thy1 primer set 2. Peak at Chromosome 9 shows endogenous Thy1 (black circle) and peak at Chromosome 1 shows inserted sequence (red circle). (D) Regional coverage of the TYW3 insertion site on Chromosome 1 using both sets of primers, showing a 419 kb deletion and spanning 5 Mb. (E) Schematic of the inserted sequence on Chromosome 1. The transgene was inserted multiple times between Lgsn and Khdrbs2 in Chromosome 1, in the 5′ to 3′ orientation. Primers were designed to confirm the left junction between the transgene and Chromosome 1, as well as the predicted deletion. PCR product from homozygous TYW3 and C57BL/6J animals is denoted as “Tg/Tg” and “WT” respectively. (F) Primers 8 and 9 were used to confirm the junction between the 5′ end of the transgene and the 3′ end of Chromosome 1 in homozygous animals. This 292 bp band is absent in wild-type animals (G) Primers 10 and 11 were used to confirm the deletion engendered by the transgene. The 173 bp band is absent in putative homozygous TYW3 animals.