Eya4^{TgTn(Prm1-sb10,sb-Tyr)1739AOve}

A Sleeping Beauty (SB) transposon transgenic approach using co-injection of the transposon vector pT-Tybs-3'E and the SB-expressing transgene Prm-SB10. The pT-Tybs-3'E transposon vector used here has an inverted repeat/direct repeat sequence (IR/DR; the SB transposon recognition site [280 bp]), mouse tyrosinase enhancer sequence (3.65 kbp), Tyro minigene (TyBS; 4.1 kbp), and right IR/DR (280 bp). The mouse tyrosinase enhancer sequence is from the region positioned ~14 kbp upstream from exon 1 in the mouse genome. The tyrosinase minigene (TyBS) contains the tyrosinase upstream regulatory sequences (2.1 kbp from the BALB/c tyrosinase promoter and the first 65 bp of exon 1), followed by the C57BL/6-derived Tyrs-J cDNA sequence (1.9 kbp including cysteine at amino acid 103, glycine at amino acid 346, and the stop codon with polyA signal). The donating investigator did not report whether the IR/DR sequences are outward-facing (pointed away from the Tyro sequences) or inward-facing (pointed toward the Tyro sequences). The Prm-SB10 transgene used here (prm-SB10-rabbit β-globin plasmid #1611) has the mouse protamine 1 promoter (652 bp), a 25 bp linker, SB10 gene (1023 bp), and rabbit β-globin splice/polyA sequence (1170 bp). Inverse PCR analysis identified that the integration site of the co-injected transgenes is on chr10 A3(+):22,797,990 bp [NCB137/mm9]: ~25 kbp away from Eya4 (22,823,974-23,069,701 [NCB137/mm9]). Although the OVE1739A mice still carry at least one copy of the SB10 transgene, there is no evidence of remobilization of the transposon. RT-PCR confirmed the absence of transcript expression. (J:175597)