B6.129-Tlr4^tm1.1Jke/J

Stock number: 036079
Product name: Tlr4^fl

Research areas: Cancer Research, Diabetes and Obesity Research, Immunology, Inflammation and Autoimmunity Research, Research Tools

Description

Tlr4^fl mice have loxP sites flanking exons 2-3 of the toll-like receptor 4 gene. Exposure to Cre recombinase removes the floxed sequence - creating a null allele. These Tlr4^fl mice may be useful in studying diabetes, insulin resistance, and innate immune responses.

Detailed description

The Tlr4 gene encodes a protein that has a critical role in pathogen recognition and activation of the innate immune system, and is a key mediator of pro-inflammatory responses.

The Tlr4^fl allele has loxP sites flanking exons 2-3 of the Tlr4 gene. Prior to introduction of Cre recombinase, mice heterozygous or homozygous for the Tlr4^fl allele are viable and fertile with normal breeding, and have no gross physical or behavioral abnormalities.

When Tlr4^fl are bred to mice that express Cre recombinase, the resulting offspring will have exons 2-3 deleted - creating a null allele in the cells/tissues as determined by the cre promoter.
For example, when bred to germline Cre-expressing mice (Zp3-Cre; Stock No. 003651), the resulting Tlr4 global knock-out homozygotes were confirmed to have no Tlr4 mRNA expression in hepatocytes (by qPCR). When those Tlr4 global knock-out homozygotes were treated with LPS, they exhibited a >90% reduction in plasma levels of Tnfα.
Furthermore, when Tlr4^fl mice are bred to Albumin-Cre transgenic mice (such as Stock No. 003574), the resulting liver-specific knock-out mice also exhibit a ~25% reduction in plasma levels of Tnfα and had lower non-fasted blood glucose levels when fed a high-fat diet, improved glucose tolerance, and enhanced insulin sensitivity.
Moreover, when crossed to LysM-Cre mice (such as Stock No. 004781), the resulting myeloid cell-specific knock-out mice have dramatically reduced (>90%) mRNA expression of Tlr4 in the peritoneal macrophages. In addition, these myeloid cell-specific knock-out mice show >35% reduction of circulating Tnfα in response to LPS challenge (Jia et al., 2014).

Of note, The Jackson Laboratory Repository also offers a similar Tlr4 floxed allele (Stock No. 024872) which has only exon 3 flanked by loxP sites. Those Tlr4 floxed mice may carry genetic contributions from both C57BL/6N and C57BL/6J, while Stock No. 036079 has both exons 2 and 3 flanked by loxP sites and has been backcrossed at least 10 generations to C57BL/6J.

Development

The Tlr4 floxed allele (also called Tlr4^fl) was created in the laboratory of Dr. Joel Elmquist (UT Southwestern Medical Center). First, a targeting vector was designed to insert a loxP site upstream of exon 2, and a second loxP site followed by an FRT-flanked PGK-neo cassette just downstream of exon 3 of the toll-like receptor 4 locus (Tlr4) on chromosome 4. The construct was electroporated into 129/SvJ-derived embryonic stem (ES) cells. Correctly targeted ES cells were injected into recipient blastocysts, and chimeric mice were bred with C57BL/6J for germline transmission. Offspring were bred with Flpe-expressing mice (ROSA26-FLPe on a 129S4/SvJaeSor background; Stock No. 003946) for germline deletion of the PGK-neo cassette. The resulting Tlr4^fl colony was then backcrossed to C57BL/6J mice for at least 10 generations (and the Flpe-expressing allele was removed) prior to sending males to The Jackson Laboratory Repository in 2021. 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).

Allele

Symbol: Tlr4^tm1.1Jke
Name: targeted mutation 1.1, Joel Elmquist
MGI accession ID: MGI:6511933
Generation method: Targeted
Attributes: Conditional ready (e.g. floxed)
Synonyms: Tlr4^fl
Marker symbol: Tlr4
Marker name: toll-like receptor 4
Marker synonyms: ARMD10; CD284; lipopolysaccharide response; Lps; Rasl2-8; TLR-4; TOLL
Chromosome: 4
Strain of origin: 129
Molecular note: The targeting vector is designed to insert a loxP site upstream of exon 2, and a second loxP site followed by an FRT-flanked PGK-neo cassette just downstream of exon 3. Flp-mediated recombination removed the FRT-flanked neo.