Igs7^{tm228(tetO-GCaMP6m,CAG-ChRmine*/oScarlet*,CAG-tTA2)Tasic}

The targeting vector has a Tet response element (tetO), loxP-STOP-loxP cassette, GCaMP6m, a CAG promoter, Lox2272-STOP-Lox2272 cassette, ChRmine*/oScarlet* sequence, and then a CAG promoter, LoxN-STOP-LoxN cassette, and IRES-tTA2 sequence - all inserted into the T1 TIGRE locus (an intergenic region on chromosome 9 [between AB124611 and Carm1] determined to have low basal transcriptional activity and allow high Cre and/or Tet inducibility). ). ChRmine is a pump-like cation-conducting channelrhodopsin, derived from Rhodomonas lens, that exhibits extremely high current and light sensitivity, as well as a markedly red-shifted spectrum. This is fused with a membrane trafficking signal (TS) and oScarlet fluorescent protein, which is based on the sequence of mScarlet with an additional E95D mutation to increase neuronal cell trafficking. GCaMP6M is an ultrasensitive EGFP fluorescent calcium detector of neuronal activity with medium decay and response kinetics. Of note, the allele was designed with an IRES upstream of the tTA2 to reduce/eliminate leakiness in the absence of Cre recombinase. (J:101977)