Tmprss2^{em2.1(TMPRSS2)Synbl}

Cas9 and gRNA Tmprss2-crRNA_5 (protospacer sequence 5-GTGCTCAGACTGATACCCG-3) are used to introduce the following sequences at codon 19 in exon 3 of the mouse Tmprss2 gene: GGSG linker, P2A, human TMPRSS2 cDNA coding sequence with SV40, 1837 bp human TMPRSS2 3UTR, 112 bp human intergenic sequence immediately 3 of TMPRSS2 polyadenylation region, cassette comprised of a mouse U6 promoter-driven Tmprss2-gRNA5 guide RNA with 2xPolIII terminator, all in reverse orientation relative to Tmprss2 gene, splice acceptor and SV40 polyadenylation sequence designed to trap and truncate any mouse Tmprss2 isoforms that may splice past the hTMPRSS2 cassette. FRT-flanked PGK neomycin cassette with a bGHpA. The FRT sites allow Flp-mediated removal of the neomycin cassette from targeted embryonic stem cell clones as well as collapse of targeted tandem integration events if required. Flp-mediated recombination removed the FRT-flanked neo cassette. (J:101977)