Igs7^{tm92.1(tetO-ECFP*/Venus*)Hze}

The vector was designed with (from 5' to 3') an FRT3 site, two copies of chicken beta-globin HS4 insulator element, a modified Tet response element promoter, a floxed-STOP cassette, the YCX2.60 coding sequence [an enhanced cyan fluorescent protein with C-terminal 11 amino acids deleted (ECFPDeltaC11 with R27K and K213N mutations, a rat calmodulin DNA fragment (CaM; aa 5-148 with N60D and D78Y mutations [to increase the fluorescence change for small calcium transients] and E127V mutation), a GG linker, a chicken smooth muscle M13 fragment of myosin light chain kinase (with I-to-S mutation in the 20th aa of the sequence), and a circularly permutated Venus enhanced yellow fluorescent protein (cp173 Venus; aa 174-239, a 15 bp linker and aa 1-173)], a WPRE sequence, a BGH polyA, two copies of chicken beta-globin HS4 insulator element, an AttB site, a PGK-5'hygro cassette, an RNA splice donor and a FRT5 site. The fluorescence resonance energy transfer (FRET)-based calcium sensor Yellow Cameleon variant X 2.60 (YCX2.60) is an improved version of YC2.60 with expanded dynamic range and 2-3-fold greater dissociation constant. The fusion gene is an enhanced cyan fluorescent protein with C-terminal 11 amino acids deleted, a rat calmodulin DNA fragment (CaM; aa 5-148 with N60D and D78Y mutations [to increase the fluorescence change for small calcium transients] and E127V mutation), a GG linker, a chicken smooth muscle M13 fragment of myosin light chain kinase (with I-to-S mutation in the 20th aa of the sequence), and a circularly permutated Venus enhanced yellow fluorescent protein (cp173 Venus; aa 174-239, a 15 bp linker and aa 1-173). PhiC31-mediated recombination removed the AttB/AttP-flanked PGK-hygromycin-SV40polyA cassette and replaced it with the recombined AttB/AttP site (AttL). (J:219930)