C57BL/6N-Fgd5^{tm3(cre/ERT2)Djr}/J

Stock number: 027789
Product name: Fgd5^{ZsGr•CreERT2}
Strain synonyms: Fgd5-CreERT2
Research areas: Hematological Research, Research Tools

Description

These Fgd5-CreERT2 knock-in/knock-out mice express ZsGreen and tamoxifen inducible Cre recombinase from the endogenous promoter/enhancer elements of the Fgd5 locus. When induced, cre activity is observed in hematopoietic stem cells and endothelial cells. This strain also serves as a reporter strain, and may be suitable for identification and purification of hematopoietic stem cells based on single-color fluorescence.

Detailed description

The Fgd5 gene encodes a protein that upregulates CDC42 activity and is a marker for immunophenotypic hematopoietic stem cells. These Fgd5^{ZsGr•CreERT2} mice carry a knock-in/knock-out mutation of the Fgd5 gene in which ZsGreen and creERT2 cassettes (separated by a 2A peptide cassette) have been inserted in frame into the first coding exon. Under control of the endogenous upstream promoter/enhancer elements, tamoxifen-inducible cre activity is observed in ZsGreen+ LSKCD150+ hematopoietic stem cells. Homozygotes are not viable, homozygous embryos die around E11.5–E12.0.

The Cre-ERT2 fusion protein consists of Cre recombinase fused to a 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, Cre-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.

Development

A targeting vector was used to insert ZsGreen and creERT2 cassettes, separated by a 2A peptide cassette, in frame into exon 1 with a FRT site flanked PGK-NEO cassette downstream of the ZsGreen cassettes. The construct was electroporated into C57BL/6NTac derived B6-3 embryonic stem (ES) cells. Correctly targeted ES cells were injected into blastocysts. The resulting chimeric animals were tested for germline transmission. Upon arrival at The Jackson Laboratory, the mice were crossed to C57BL/6NJ (Stock No. 005304) at least once to establish the colony.

Allele

Symbol: Fgd5^{tm3(cre/ERT2)Djr}
Name: targeted mutation 3, Derrick Rossi
MGI accession ID: MGI:5642327
Generation method: Targeted
Attributes: Recombinase-expressing; Reporter; Inducible
Marker symbol: Fgd5
Marker name: FYVE, RhoGEF and PH domain containing 5
Chromosome: 6
Strain of origin: C57BL/6NTac
Expressed genes: GFP,Green Fluorescent Protein,; cre/ERT2,Cre recombinase and estrogen receptor 1 (human) fusion gene,
Site of expression: When induced, cre activity is observed in hematopoietic stem cells and endothelial cells.
Molecular note: The targeting vector consists of ZsGreen and creERT2 cassettes separated by a 2A peptide cassette and followed by an FRT-flanked neomycin cassette inserted in frame into exon 1.