These floxed mutant mice possess loxP sites flanking exon 7 of the Traf6 gene and a STOP cassette, with a downstream IRES-YFP. Cre recombinase-mediated excision of exon 7 and the STOP cassette generates a null allele and allows expression of the YFP from the endogenous Traf6 locus. This strain may be useful for generating conditional mutations in applications related to signal transduction, autophagy regulation, TRAF pathway and toll-like receptor (TLR) pathway.
Important Note: retinal degeneration mutant allele Crb1rd8 discovered in the frozen bankstock for Stock No. 030849. See the Strain Development section for more information.
Yongwon Choi, University of Pennsylvania Perelman School of Medicine
The targeted Traf6 gene encodes a signal transduction protein that is part of the NF-kappa-B pathway and has been implicated in the pathogenesis of Parkinson disease. These mice possess loxP sites on either side of exon 7 of the targeted gene as well as a STOP cassette. Exon 7 encodes the last zinc finger domain. Mice that are homozygous for this allele are viable and fertile. When these mutant mice are bred to mice that express Cre recombinase, resulting offspring will have exon 7 and STOP cassette, deleted in the cre-expressing tissues and express EYFP from the endogenous Traf6 promoter. Removal of the floxed sequence creates a null EYFP reporter allele.
A targeting vector containing a loxP site flanked NEO selection cassette was utilized in the construction of this mutant. This selection cassette was inserted upstream of exon 7 of the targeted gene, and a STOP cassette, a loxP site, and an IRES-EYFP cassette was inserted downstream of exon 7. This construct was electroporated into unspecified embryonic stem (ES) cells. Correctly targeted ES cells were injected into C57BL/6 blastocysts. The resulting chimeric animals were tested for germline transmission. Cre mediated recombination removed the loxP site flanked NEO selection cassette. The mice were backcrossed to C57BL/6 for approximately 12 generations. Upon arrival, sperm was cryopreserved. To establish our live colony, an aliquot of frozen sperm was used to fertilize C57BL/6J oocytes (Stock No. 000664).
Important Note: retinal degeneration mutant allele Crb1rd8 discovered in the frozen bankstock for Stock No. 030849:
A 48 SNP (single nucleotide polymorphism) panel analysis, with 43 markers covering all 19 chromosomes and the X chromosome, as well as 5 markers that distinguish between the C57BL/6J (B6J) and C57BL/6N (B6N) substrains, was performed on the males sent to The Jackson Laboratory Repository (and is the source of frozen sperm used for cryorecovery since 2020). 42 of 43 markers throughout the genome suggested a C57BL/6 genetic background (see additional data below). For the 5 markers that determine B6J from B6N, 4 were found to have C57BL/6 substrain contributions: homozygous B6N SNP on chromosome 1 ~139 Mbp (see more below), heterozygous B6N/B6J SNP on chromosome 8 [~58 Mbp], heterozygous B6N/B6J SNP on chromosome 15 [~57 Mbp] and either homozygous B6N SNP or homozygous B6J SNP on chromosome 19 [~50 Mbp]).
The homozygous B6N SNP on chromosome 1 ~139 Mbp indicates they have the recessive retinal degeneration mutant allele Crb1rd8. Researchers can choose cryorecovery via IVF with C57BL/6J oocytes to receive mice heterozygous for that recessive allele. Of note, the mice were homozygous C57BL/6 SNP on chromosome 5 ~108 Mbp (Pde6b), indicating they do not have the retinal degeneration mutant allele Pde6brd1.
Additionally, there appears to be a genetic background contribution other than C57BL/6 on chromosome 1 ~87 Mbp.
|Allele Name||targeted mutation 2.1, Yongwon Choi|
|Allele Type||Targeted (Conditional ready (e.g. floxed), No functional change)|
|Gene Symbol and Name||Traf6, TNF receptor-associated factor 6|
|Strain of Origin||Not Specified|
|General Note||The version of this allele that contains the neomycin selection cassette, Traf6tm2Ywc, has been analyzed.|
|Molecular Note||A floxed neomycin selection cassette was placed upstream of exon 7 and a STOP cassette followed by a loxP site and an YFP reporter were placed downstream of exon 7. Cre-mediated recombination removed the floxed neo cassette.|
When maintaining a live colony, these mice can be bred as homozygotes.
When using the Traf6floxed mouse strain in a publication, please cite the originating article(s) and include JAX stock #030849 in your Materials and Methods section.