STOCK Isl1^{tm1(cre/Esr1*)Krc}/SevJ

Stock number: 029566
Product name: isl1-MerCreMer
Research areas: Developmental Biology Research, Research Tools

Description

These Isl1 MerCreMer knock-in mice express a tamoxifen-inducible cre recombinase from the endogenous promoter/enhancer elements of the Isl1 gene. When induced, cre activity is observed in Isl1 expressing cells in embryos. These mice may be useful for generating conditional mutations for studying cell lineages during development of the nervous system, heart, pituitary gland, pancreas and stomach.

Detailed description

These targeted mutation knock-in mice have a tamoxifen inducible Cre-mediated recombination system ("MerCreMer") driven by the endogenous Isl1 (ISL1 transcription factor, LIM/homeodomain) promoter. Inducible Cre mediated recombination occurs in Isl1 expressing cells in embryos. Tamoxifen treatment at E7.5 results in Cre recombinase activity at E9.5 in cardiac progenitor cells. Mice that are heterozygous for the targeted mutation are viable and fertile. Homozygotes have an embryonic lethal phenotype.

Of note, this double fusion protein has substantially greater Cre recombinase activity with less promiscuity compared with a single CreMer fusion protein. As the cre is flanked on each end with a mutated murine estrogen receptor ligand binding domain (amino acids 281-599, G525R); Cre expression is tamoxifen inducible yet estrogen insensitive.

The cre/Esr1* fusion protein consists of Cre recombinase fused to a G525R mutant form of the mouse 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, cre/Esr1* 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.

Development

A targeting vector containing IRES-MerCreMer sequence and an FRT-site flanked NEO cassette (FRT-mclNeo) was used to disrupt part of exon 1, including the ATG codon, exon 2, and part of exon 3, of the targeted gene. The construct was electroporated into unspecified 129 derived embryonic stem (ES) cells. Correctly targeted ES cells were injected into blastocysts. The resulting male chimeric animals were crossed to C57BL/6J females. The mice were crossed to a FLP recombinase expressing line to remove the FRT-site flanked NEO cassette. The mice were then crossed to Black Swiss outbred mice for 5 generations. Upon arrival, sperm was cryopreserved. To establish our live colony, an aliquot of frozen sperm was used to fertilize C57BL/6J oocytes (Stock No. 000664).

Allele

Symbol: Isl1^{tm1(cre/Esr1*)Krc}
Name: targeted mutation 1, Kenneth R Chien
MGI accession ID: MGI:3530632
Generation method: Targeted
Attributes: Recombinase-expressing; Inducible
Synonyms: Isl1^{tm1(cre/Esr1)Krc}; isl1-MCM; isl1-MerCreMer; isl1-mER-Cre-mER; MerCreMer
Marker symbol: Isl1
Marker name: ISL1 transcription factor, LIM/homeodomain
Marker synonyms: Isl-1; isl-1=homeobox; Islet 1; ISLET1
Chromosome: 13
Strain of origin: 129
Expressed genes: cre/Esr1,Cre recombinase and estrogen receptor 1 fusion gene,
Site of expression: When induced by tamoxifen, cre activity is observed in Isl1 expressing cells in embryos.
Molecular note: A targeting cassette containing a tamoxifen-dependent cre flanked by two mutated mouse estrogen receptors that sequester cre to the cytoplasm, followed by an FRT-flanked neo was inserted into the Isl1 locus by homologous recombination. Flp-mediated recombination removed the FRT-flanked neo cassette. A short segment of coding region containing the endogenous ATG codon was deleted.