B6.Cg-Tg(Dmp1-cre/ERT2)0022Pdp/IkalJ

Stock number: 032757
Product name: 10kb-Dmp1-Cre-ERT2
Research areas: Cell Biology Research, Developmental Biology Research, Internal/Organ Research, Research Tools

Description

10kb-Dmp1-Cre-ERT2 transgenic line 0022 has the 10 kbp promoter/enhancer sequences from the mouse dentin matrix protein 1 locus (Dmp1) directing tamoxifen-inducible Cre recombinase (cre/ERT2) expression to osteocytes, some mature osteoblasts and rare bone lining cells.

Detailed description

Dentin matrix protein 1 (encoded by DMP1) is an extracellular matrix protein critical for proper mineralization of bone and dentin, is present in diverse bone/tooth cell populations and functions in osteoblast maturation by coordinating mineralized matrix formation.

The 10kb-Dmp1-Cre-ERT2 transgene has the ~10 kbp mouse Dmp1 promoter/enhancer regions directing tamoxifen-inducible Cre recombinase expression. CreER^T2 fusion gene activity is inducible; observed following tamoxifen administration. When 10kb-Dmp1-Cre-ERT2 transgenic mice are bred with mice containing lox-flanked (floxed) sequences, tamoxifen-inducible Cre-mediated recombination will result in deletion of floxed sequences in Dmp1-expressing cells in the offspring. Hemizygous and homozygous mice are viable and fertile with no reported phenotype abnormalities.

Specifically, tamoxifen-inducible Cre recombinase activity recapitulates the endogenous Dmp1 expression pattern. The donating investigator reports that in founder line 0022, when combined with mice carrying Gt(ROSA)26Sor^{tm9(CAG-tdTomato)Hze} (Ai9), progeny exhibit some leakiness in about 20% of osteocytes prior to tamoxifen administration. Following tamoxifen administration, tdTomato is observed in osteocytes, osteoblasts, rare bone lining cells, and some cells within the bone marrow, and some cells within the muscle. Dr. Kalajzic reports characterizing mouse line 0022 in the following publications: PMID:23072918, PMID:27507737, PMID:32053258.

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

The 10kb-Dmp1-Cre-ERT2 transgenic mice (also called ^10kbDmp1-CreERt2) were created in the laboratory of Dr. Paola Divieti-Pajevic (Massachusetts General Hospital). First, the ~14 kbp fragment (-9624 to +4439) of the mouse dentin matrix protein 1 locus (Dmp1) containing a 9624 bp promoter region, the 95 bp exon 1, a 4326 bp intron 1 and a 17 bp of initial non-coding region of exon 2 was obtained. This promoter/enhancer region was placed upstream of a cDNA sequence encoding the CreER^T2 fusion gene (Cre recombinase fused to a G400V/M543A/L544A triple mutation of the human estrogen receptor ligand binding domain), followed by an SV40 polyadenylation signal. The resulting transgene (~16 kbp) was microinjected into the pronucleus of B6C3F1/Tac hybrid mice. Founder mice were bred to B6C3F1/Tac wildtype mice for germline transmission. Founder line 0022 was identified to have tamoxifen-inducible Cre recombinase activity in both osteocytes and mature osteoblasts. Mice from founder line 022 were backcrossed to C57BL/6 for at least 5 generations. Mice from the colony were transferred into the laboratory of Dr. Ivo Kalajzic (University of Connecticut) in 2012 and combined with B6.Cg-Tg(Col1a1*2.3-GFP)1Rowe/J (Stock No. 013134) mice. The colony was maintained for many generations prior to sending hemizygous male mice with black coat color to The Jackson Laboratory Repository. To establish our live colony, sperm from those males was used to fertilize C57BL/6J oocytes (Stock No. 000664). The Tg(Col1a1*2.3-GFP)1Rowe allele was bred out of our colony.
Of note, the donating investigator reports that, at least once during backcrossing, a hemizygous female was bred to a C57BL/6 inbred male (thus the Y chromosome of the congenic strain is of C57BL/6 origin). The chromosome location of transgene insertion is not known.

Allele

Symbol: Tg(Dmp1-cre/ERT2)0022Pdp
Name: transgene insertion 0022, Paola Divieti Pajevic
MGI accession ID: MGI:5792965
Generation method: Transgenic
Attributes: Recombinase-expressing; Inducible
Synonyms: 10kb-Dmp1-Cre-ERT2; Tg(Dmp1-cre/ERT2)1Pdp
Marker symbol: Tg(Dmp1-cre/ERT2)0022Pdp
Marker name: transgene insertion 0022, Paola Divieti Pajevic
Chromosome: UN
Strain of origin: (C57BL/6 x C3H)F1
Molecular note: A fragment (-9624 - +4439) of the mouse Dmp1 gene containing a 9624 bp promoter region, the 95 bp exon 1, 4326 bp intron 1 and 17 bp of initial non-coding region of exon 2 was cloned into a vector in front of the cre/ERT2 cDNA. The linearized vector was injected into B6C3F1 pronuclei. 14 pups that were positive for cre integration were identified and six independent founders were crossed to wild-type B6 mice. Two founders demonstrated penetrant excision of the STOP cassette when crossed to Rosa26-lacZ reporter mice, and were analyzed in greater detail. The two transgenic founder lines (D77 and 0022) were identified to have tamoxifen-inducible Cre recombinase activity in both osteocytes and mature osteoblasts. The two lines were similar with the only difference that line D77 had greater expression in mature osteoblasts.