Tg(Gja5-GCaMP3*/mCherry)B15-2Mik

The transgenic construct is made using a C57BL/6J mouse bacterial artificial chromosome (BAC) RP24-255O4 containing the entire Gja5 locus as well as ~85 kbp of 5' flanking sequences and ~61 kbp of 3' flanking sequences (including the partially overlapping 4930573H18Rik locus). Using homologous recombination/BAC recombineering, a 3006 bp GCaMP-GR-pA construct is inserted into the ATG start site of the BAC Gja5 gene (replacing nucleotides aagatggg at the initiation codon of Gja5 in exon 2). No other loci on the BAC were altered. The 13.5 kbp BAC vector (pTARBAC1) contained a chloramphenicol selection cassette. GCaMP-GR is a calcium-sensing molecule composed of a 34 aa polyHis plasmid leader sequence, a 13 residue peptide of chicken smooth muscle myosin light chain kinase, a circularly permutated EGFP, a rat calmodulin DNA fragment, a flexible alanine-proline repeat linker and the mCherry fluorescent protein. Founder line B15-2 has high sensor expression in arterial endothelial cells, atrial myocytes and cardiac Purkinje fibers. (J:272385)