Ple287-icre/ERT2;mEMS6107 mice have the Ple287-icre/ERT2 transgene targeted as a single copy "knock-in" into the upstream region of the hypoxanthine guanine phosphoribosyl transferase 1 (Hprt1) locus on the X Chromosome. Heterozygous females and hemizygous males are viable and fertile, with the promoter/enhancer/regulatory regions of the human sprouty homolog 1 (SPRY1) locus directing expression of the tamoxifen-inducible, improved Cre recombinase (icre/ERT2). The donating investigator reports RT PCR expression as "Positive (Hippocampus, Telencephalon, Olfactory Bulb)." The phenotype of homozygous mice has not been evaluated to date (January 2014).

The iCre/ER^T2 fusion protein used here consists of a codon-improved Cre recombinase fused to a G400V/M543A/L544A triple mutant form of the human estrogen receptor which does not bind its natural ligand (17β-estradiol) at physiological concentrations but will bind the synthetic estrogen receptor ligands 4-hydroxytamoxifen (OHT or tamoxifen) and, with lesser sensitivity, ICI 182780. Restricted to the cytoplasm, iCre/ERT2 can only gain access to the nuclear compartment after exposure to tamoxifen. To counteract the mixed estrogen agonist effects of tamoxifen injections, which can result in late fetal abortions in pregnant mice, progesterone may be coadministered.

Ple287-icre/ERT2;mEMS6107 mice were created by the CanEuCre and Pleiades Promoter Project (Centre for Molecular Medicine and Therapeutics, University of British Columbia). These mice have the Ple287-icre/ERT2 transgene targeted as a single copy "knock-in" into the upstream region of the hypoxanthine guanine phosphoribosyl transferase 1 (Hprt1) locus on the X Chromosome. As such, the promoter/enhancer/regulatory regions of the human sprouty homolog 1 (SPRY1) locus are designed to direct expression of the tamoxifen-inducible, improved Cre recombinase (icre/ERT2). More specific details are described below.

The RP11-1126.D8 human bacterial artificial chromosome (BAC) was obtained and retrofitted to contain a human HPRT1 complementary sequence (containing exon 1, intron 1, exon 2 and part of intron 2), a mouse 3' Hprt1 homology arm, an I-Sce linearization cut site, and a mouse 5' Hprt1 homology arm; all inserted into the sacB gene in the BAC vector backbone (thus disrupting the sacB gene and removing the loxP site in the vector backbone). Next, an iCre/ERT2 fusion gene and an Ffrt-flanked kanamycin cassette were inserted directly at the ATG of the human sprouty homolog 1 (SPRY1) locus. The kanamycin cassette was then permanently removed via transient transfection of a FLP recombinase plasmid; creating the Ple287-icre/ERT2 BAC (bEMS248) used as the source of the Ple287-icre/ERT2 transgene. This transgene was targeted as a single copy knock-in to the Hprt1b-m3 mutant locus on the X Chromosome via electroporation into mEMS4857.02 embryonic stem cells (described below). The resulting chimeric males (founder line mEMS6107) were bred to C57BL/6J females to establish the mutant colony. The donating investigator reports that Ple287-icre/ERT2;mEMS6107 mutant animals were backcrossed to C57BL/6J mice for 2 generations, and then sperm was frozen from black hemizygous males at the University of British Columbia. In 2013-2014, frozen sperm was sent to the Mutant Mouse Regional Resource Center at The Jackson Laboratory (MMRRC-JAX). When rederiving the live colony, an aliquot of frozen sperm is used to fertilize oocytes from C57BL/6J female mice (Stock No. 000664); the resulting obligate heterozygous females are bred at least one generation to C57BL/6J mice to establish the live colony.

mEMS4857.02 embryonic stem (ES) cells are a male ES cell line made from an NE4.75F4 blastocyst from C57BL/6NTac-Aw-j/Aw-j Hprt1b-m3/Y mice (created by four generations of breeding C57BL/6NTac-Aw-J together with C57BL/6NTac-Hprt1b-m3). The parental lines were individually made by backcrossing C57BL/6J-Aw-J/J (Stock No. 000051) four generations onto C57BL/6NTac, and backcrossing B6.129P2-Hprt1b-m3/J (Stock No. 002171) six generations onto C57BL/6NTac, respectively.

The MMRRC Centers have developed a genetic QC pipeline using MiniMUGA array genotyping to provide additional information on strain backgrounds for MMRRC congenic and inbred strains. For more information on when data may be available, or to request genotyping for a strain of interest, please contact csmmrrc@jax.org. Older strains may not have this information.

Simpson [Ph.D.], Elizabeth M.