This allele has three membrane-targeted fluorescent protein sequences - each uniquely flanked with pairs of incompatible lox sites (in the same direction) inserted between exons 1 and 2 of the Gt(ROSA)26Sor locus with a CAG hybrid promoter/enhancer. The targeting vector is designed with (from 5' to 3') two copies of chicken beta-globin HS4 insulator element (2xINS; increase downstream gene expression and minimize regulatory interference), a CMV-IE enhancer/chicken beta-actin/rabbit beta-globin hybrid promoter (from pCAGGS), a loxN site, a lox2272 site, a loxP site, the non-cytotoxic/non-fluorescent Phialidium yellow fluorescent protein variant PhiYFP with nuclear localization signal (N-PhiMut), SV40 polyA signal sequence, a second loxN site, a full consensus KOZAK sequence, the Mb2-YFP gene (monomeric enhanced yellow fluorescent protein [mYFPA206K] with N-terminal 41 amino acid membrane-targeting tag sequence), in-frame Thosea asigna virus 2A cleavage site (T2A; mediates ribosomal skipping), a woodchuck hepatitis virus post-transcriptional regulatory element (WPRE; to enhance mRNA transcript stability), SV40 polyA signal sequence, a second lox2272 site, a full consensus KOZAK sequence, the Mb2-Tomato gene (non-oligomerizing tandem-dimer DsRed fluorescent protein variant tdTomato with N-terminal 41 amino acid membrane-targeting tag sequence), in-frame T2A sequence, WPRE sequence, SV40 polyA signal sequence, a second loxP site, a full consensus KOZAK sequence, the Mb2-HA-TFP1 gene (monomeric teal fluorescent protein mTFP1 with N-terminal hemagglutinin tag and 41 amino acid membrane-targeting tag sequence), in-frame T2A sequence, WPRE sequence, SV40 polyA signal sequence, an FRT site flanked neomycin resistance gene cassette, and a second 2xINS sequence. (J:252386)
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