B6.Cg-Dcpp1^{tm1.1(cre/ERT2)Ovi}/J

Stock number: 028731
Product name: Dcpp1^GCE
Research areas: Endocrine Deficiency Research, Reproductive Biology Research, Research Tools

Description

Dcpp1^GCE knock-in mice express a EGFP/cre/ERT2 fusion protein in serous demilune cells of the sublingual gland and intercalated duct cells of the parotid gland. These mice may have applications in studies requiring conditional gene manipulation or cell lineage tracing in sublingual and parotid glands.

Detailed description

Demilune cell and parotid protein 1, encoded by the Dcpp1 gene, is a secretory protein found in oviduct epithelium and salivary glands. These knock-in mice express the GCE cassette (an EGFP/creERT2 fusion gene) from the endogenous Dcpp1 locus. When Dcpp1^GCE mice are bred with mice containing loxP-flanked sequence, tamoxifen-inducible, cre-mediated recombination will result in deletion of the floxed sequences. Tamoxifen administration in double mutant mice, carrying this GCE cassette and a reporter, induces Cre recombination in serous demilune cells of the sublingual gland and intercalated duct cells of the parotid gland. The inducible cre recombinase activity pattern mimics the endogenous Dcpp1 expression pattern. EGFP expression is not detected by direct fluorescence or anti-GFP antibody (immunofluorescence). Although homozygotes are predicted to be viable and fertile, the Donating Investigator has not made the strain homozygous. During backcrossing, the Y chromosome may not have been fixed to the C57BL/6J genetic background.

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 construct containing the GCE cassette (EGFP and Cre/ERT2 fusion gene) followed by SV40 polyadenylation sequence and an FRT-flanked neo cassette was used to disrupt exon 2 of the targeted gene. The construct was electroporated into (129S6/SvEvTac x C57BL/6NCrl)F1-derived G4 embryonic stem (ES) cells. Correctly targeted ES cells were injected into C57BL/6J blastocysts, and chimeras were bred to C57BL/6J mice. The resulting Dcpp1-GCE-Neo allele heterozygotes were then bred to Actin-Flippase mouse strain on a B6.129S4 genetic background (Stock No. 009086) to remove the FRT-flanked NEO cassette, generating Dcpp1^GCE offspring. The mice were then backcrossed to C57BL/6 for 3 generations. During backcrossing, the Y chromosome may not have been fixed to the C57BL/6J genetic background. Upon arrival, B6N3 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: Dcpp1^{tm1.1(cre/ERT2)Ovi}
Name: targeted mutation 1.1, Catherine E Ovitt
MGI accession ID: MGI:5661581
Generation method: Targeted
Attributes: Recombinase-expressing; Inducible
Marker symbol: Dcpp1
Marker name: demilune cell and parotid protein 1
Chromosome: 17
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: The fusion cre is expressed in intercalated duct cells of parotid gland and serous cells of sublingual gland.
Molecular note: The EGFP-cre/ERT2 (GCE) fragment with an SV40 poly adenylation site and neomycin resistance cassette were inserted immediately downstream of the translational initiation codon ATG. The knock-in construct interrupted coding sequence in exon 2 and placed the GCE gene under the control of the endogenouse regulatory sequences. Flp-mediated recombination removed the selection cassette. No expression of GFP was detected with fluorescence or using antibody to GFP. The fusion cre is expressed in intercalated duct cells of parotid gland and serous cells of sublingual gland.
General note: No evidence of ectopic expression in lungs, kidney, pancreas, or ovaries.