STOCK Tmem178b^{tm1c(KOMP)Wtsi} Tmem178^tm1.1Sud/J

Stock number: 036906
Product name: Tmem178ab cKO, Tmem178a^fl;Tmem178b^fl
Research areas: Neurobiology Research, Research Tools

Description

Tmem178ab cKO mice carry conditional mutations in both the mouse Tmem178 (transmembrane protein 178; also called Tmem178a) and Tmem178b (transmembrane protein 178B) genes. Exon 4 of each gene is flanked by loxP sites in the Tmem178a^fl (also called Tmem178a cko) and Tmem178b^fl (also called Tmem178b cko) alleles. The Tmem178a^fl allele also expresses an IRES-mVenus reporter, detectable though GFP immunohistochemistry in the absence of Cre recombinase.

Detailed description

Tmem178ab cKO mice carry conditional mutations in both the mouse Tmem178 (transmembrane protein 178; also called Tmem178a) and Tmem178b (transmembrane protein 178B) genes. Exon 4 of each gene is flanked by loxP sites in the Tmem178a^fl (also called Tmem178a cko) and Tmem178b^fl (also called Tmem178b cko) alleles. The Tmem178a^fl allele also expresses an IRES-mVenus reporter, detectable though GFP immunohistochemistry in the absence of Cre recombinase. Mice homozygous for both floxed alleles are viable and fertile.

Constitutive Cre recombinase-mediated excision of the floxed regions (e.g. through lentiviruses) reduces levels of protein and disrupts the production of full-length proteins in both genes, resulting in lower body weights and impaired survival on a mixed genetic background.

Development

Tmem178a^fl (Tmem178a cko) allele: The initial construct for this allele incorporated two modified copies of mouse Tmem178 exon 4 integrated on mouse Chromosome 17. FRT and loxP2272 were introduced upstream of the lariat sequence in the first copy of exon 4 which carried a double HA (hemagglutinin) epitope in the cytoplasmic tail and a loxP2272 site in the 3' UTR. The second copy of exon 4 contains an IRES2-mVenus fluorescent reporter following the stop codon of the gene in addition to a loxP site in the 3' UTR; an Frt3 (F3) site follows the loxP site. Between the two copies of exon 4 is an Frt3-PGK promoter-EM7 promoter-Neomycin/Kanamycin-bGH poly(A) signal-FRT-loxP cassette. The mutation was created in 129S6 x C57BL/6J-derived embryonic stem cells. Resultant mice were crossed with Actb:FlpE mice (Stock No. 019100) to excise the first copy of exon 4 and other sequences between the two FRT sites. This left the second loxP-flanked IRES-mVenus copy of exon 4 intact. Resultant mice were crossed with CD-1 mice, and the FlpE allele was bred out of the stock.

Tmem178b^fl (Tmem178b cko) allele: The donating laboratory acquired C57BL/6N-A^tm1Brd Tmem178b^{tm1a(KOMP)Wtsi}/Wtsi mice from EMMA. This allele incorporates an FRT-lacZ-loxP-neomycin-FRT-loxP cassette in intron 3 and a loxP site in intron 4 of the Tmem178b gene on mouse Chromosome 6. The mutation was created in C57BL/6N-derived JM8.N4 embryonic stem (ES) cells. Mice were crossed with Actb:FlpE mice (Stock No. 019100) to excise the FRT-flanked region incorporating the lacZ and neomycin components. This left exon 4 of the targeted gene flanked by loxP sites in the Tmem178a cKO allele (Tmem178b^{tm1c(KOMP)Wtsi}). The FlpE allele was bred out of the stock.

Mice carrying the Tmem178a^fl and Tmem178b^fl alleles were intercrossed to create Tmem178ab cKO compound mutant mice on a mixed STOCK background composed of 129S6/SvEv, C57BL/6, and CD1.

Upon arrival at The Jackson Laboratory, sperm was cryopreserved. Cryorecovery/live colony development will involve C57BL/6J (Stock No. 000664).