STOCK Sema3d^{tm2.1(GFP/cre)Joe}/J

Stock number: 029270
Product name: Sema3D^GFPcre
Research areas: Cardiovascular Research, Research Tools

Description

Sema3d^GFPcre knock-in/knock-out mice express a GFPcre fusion protein under the direction of the Sema3d promoter. They may be useful for cellular fate mapping during pulmonary vein formation.

Detailed description

Sema3d^GFPcre mice express a GFPcre fusion protein from the endogenous sema domain, immunoglobulin domain (Ig), short basic domain, secreted, (semaphorin) 3D (Sema3d) promoter. The presence of GFPcre abolishes expression of Sema3d. SEMA3D is a secreted guidance molecule crucial for the normal patterning of pulmonary veins. Sema3d cellular derivatives populate the endothelial lining of the sinus venosus (SV) by E10.5. Expression is restricted, in the heart, to only proepicardial cells and migrating epicardial cells prior to E11.5, with strong epicardial expression persisting beyond E14.5. Homozygous males are mostly viable and fertile, while homozygous females do not breed. Heterozygous mice are viable and fertile. Sema3d^eGFPcre adult mice show abnormalities of pulmonary venous patterning, including massively enlarged right atria and ventricles. When these mice are bred with mice containing loxP-flanked sequence, Cre-mediated recombination will result in deletion of the floxed sequences in the Cre-expressing cells of the offspring.

Development

A targeting construct was designed to replace the initiation codon in exon 2 of the sema domain, immunoglobulin domain (Ig), short basic domain, secreted, (semaphorin) 3D (Sema3d) gene with an enhanced green fluorescent protein fused to a Cre Recombinase sequence (GFPcre), followed by a frt-flanked neomycin resistance (neo) cassette. The construct also replaced the remaining downstream sequence of exon 2. This construct was electroporated into 129S6/SvEvTac-derived TL1 embryonic stem (ES) cells. Correctly targeted ES cells were injected into blastocysts and the resulting chimeric animals were crossed to Tg(ACTFLPe)9205Dym mice on a mixed background to remove the neo selection cassette. These Sema3d^eGFPcre mice were bred together to remove the Flp-expressing transgene and were subsequently bred to C57BL/6 mice for at least 1 generation. Upon arrival, sperm was cryopreserved. To establish our live colony, an aliquot of frozen sperm was used to fertilize C57BL/6J (Stock No. 000664) oocytes.

Allele

Symbol: Sema3d^{tm2.1(GFP/cre)Joe}
Name: targeted mutation 2.1, Jonthan Epstein
MGI accession ID: MGI:5317896
Generation method: Targeted
Attributes: Recombinase-expressing
Marker symbol: Sema3d
Marker name: sema domain, immunoglobulin domain (Ig), short basic domain, secreted, (semaphorin) 3D
Chromosome: 5
Strain of origin: 129S6/SvEvTac
Expressed genes: GFP/cre,Green Fluorescent Protein/Cre recombinase fusion protein,
Site of expression: Expression is restricted in the heart to proepicardial cells and migrating epicardial cells.
Molecular note: An eGFP/cre fusion gene and an Frt-flanked neo cassette was inserted into the Sema3d locus at the ATG codon, replacing the remaining exon 2 sequence by homologous recombination in TL-1 ES cells. The neo was deleted by crossing germline mutants to Flp-expressing mice.