B6.Cg-Acan^{tm1(cre/ERT2)Crm}/J

Stock number: 019148
Product name: Agc1^{tm(IRES-CreERT2)}
Research areas: Developmental Biology Research, Internal/Organ Research, Research Tools

Description

Agc1^{tm(IRES-CreERT2)} mice express tamoxifen-inducible CreER^T2 from the aggrecan promoter. These mice may be useful for studying cartilage degenerative diseases such as osteoarthritis and degenerative disk disease.

Detailed description

The Agc1^{tm(IRES-CreERT2)} allele has a CreER^T2 sequence, and a frt-flanked neo cassette, inserted in the 3' untranslated region of the aggrecan (Acan) promoter/enhancer elements. Homozygous mice are viable and fertile. Aggrecan is an extracellular matrix protein expressed in the growth plate and articular cartilage, and fibrocartilage of the meniscus, trachea, and intervertebral disks. When Agc1^creERT2 mice are bred with mice containing loxP-flanked sequence, tamoxifen-inducible, Cre-mediated recombination will result in deletion of the floxed sequences in the aggrecan-expressing cells of the offspring.

Recent studies have shown that the insertion of the CreER^T2 cassette results in a deletion of 760 bp in the 3'UTR of the Acan gene. This results in a cre dosage-dependent reduction in body weight and body length by one month of age due to a reduction in the length of the growth plate and the thickness of articular cartilage. Due to this effect on skeletal growth, heterozygous Agc1^+/cre should be used for conditional deletion of a target gene in the cartilage tissue.

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 co-administered.

Development

A targeting vector was designed to insert an internal ribosome entry site (IRES)-CreER^T2 fusion protein, a frt-flanked neomycin (neo) selection cassette, in reverse orientation, and an SV40 polyadenylation signal, into 3' untranslated region of the aggrecan (Acan) targeted gene. This construct was electroporated into (129S6/SvEvTac x C57BL/6NCr)F1-derived G4 embryonic stem (ES) cells. Correctly targeted ES cells were injected into C57BL/6J blastocysts and chimeric males were bred with C57BL/6J females. The donating investigator reported that these Agc1^{tm(IRES-CreERT2)} mutant mice were bred to C57BL/6J mice for at least 5 generations (see SNP note below). Upon arrival at The Jackson Laboratory, mice were bred with C57BL/6J inbred mice (Stock No. 000664) for at least one generation to establish the colony.

A 32 SNP (single nucleotide polymorphism) panel analysis, with 27 markers covering all 19 chromosomes and the X chromosome, as well as 5 markers that distinguish between the C57BL/6J and C57BL/6N substrains, was performed on the rederived living colony at The Jackson Laboratory Repository. While the 27 markers throughout the genome suggested a C57BL/6 genetic background, 4 of 5 markers that determine C57BL/6J from C57BL/6N were found to be segregating. These data suggest the mice sent to The Jackson Laboratory Repository were on a mixed C57BL/6J ; C57BL/6N genetic background.

Allele

Symbol: Acan^{tm1(cre/ERT2)Crm}
Name: targeted mutation 1, Benoit de Crombrugghe
MGI accession ID: MGI:4420223
Generation method: Targeted
Attributes: Recombinase-expressing; Inducible
Marker symbol: Acan
Marker name: aggrecan
Chromosome: 7
Strain of origin: (129S6/SvEvTac x C57BL/6NCrl)F1
Expressed genes: cre/ERT2,Cre recombinase and estrogen receptor 1 (human) fusion gene,
Site of expression: When these mice are bred with mice containing loxP-flanked sequence, tamoxifen-inducible, Cre-mediated recombination will result in deletion of the floxed sequences in aggrecan-expressing cells (the growth plate and articular cartilage, fibrocartilage of the meniscus, trachea, and intervertebral disks) of the offspring.
Molecular note: IRES-cre/ERT2 and an FRT-flanked PGK-neo selection cassette (in reverse orientation to the target gene) was inserted into the 3' UTR of the Aggrecan gene by homologous recombination in G4 ES cells. The line was established by breeding to C57BL/6 mice. The neo cassette was not removed.