Strain Name:
C57BL/6J-Tlr4em1Svog/Mmucd
Stock Number:
071385-UCD
Citation ID:
RRID:MMRRC_071385-UCD
Other Names:
D298G/N397I (“TLR4-SNP” mice)

Strain Information

Tlr4
Name: toll-like receptor 4
Synonyms: lipopolysaccharide response, Lps, Rasl2-8
Type: Gene
Species: Mouse
Chromosome: 4
NCBI: 21898
Homologene: 41317
Genetic Alterations
Nucleotide substitutions in the endogenous Tlr4 locus (TLR4-SNP mice) were generated using CRISPR/Cas9-based targeting and homology-directed repair. Aspartic acid codon 298 (GAT) in exon 3 was changed to glycine (GGT) (p.298G) and asparagine codon 397 (AAC) in exon 3 to isoleucine (ATC) (p.N397I) using sgRNAs and single-stranded oligodeoxynucleotides (ssODN) templates with CRISPR/Cas9 technology. The mutations replicate the human p.D299G (c.896A>G, rs4986790) and p.T399I (c.1196C>T, rs4986791) mutations associated with LPS hyporesponsiveness and differential susceptibility to many infectious and non-infectious diseases.
ES Cell Line
NA
Phenotype

This is a knock-in mouse strain that expresses two Tlr4 SNPs that are homologous to those expressed in humans and have been associated with a variety of inflammatory/infections diseases. The primary phenotype of this mouse strain is a hyporesponsive reaction to LPS seen in humans bearing these SNPs. As described in PMID33216117, and subsequent publications, these mice are refractory to influenza infection, exhibit increased susceptibility to RSV, enhanced allergic responses, decreased response to immunization with TLR4-based adjuvants (including a defect in immunoglobulin class switching), and enhanced sensitivity to chemical-induced colitis (unpublished; supportive of studies showing that humans with these SNPs exhibit enhanced inflammatory bowel disease (IBD)).

Strain Development
C57BL/6J mice harboring nucleotide substitutions in the endogenous Tlr4 locus (TLR4-SNP mice) were generated using CRISPR/Cas9-based targeting and homology-directed repair by Cyagen Biosciences. The sgRNAs selected for mouse generation had a minimum of three mismatches located predominantly in intergenic regions. For each sgRNA, only four potential exonic off-target sites were predicted, none of which ranked in the top 40 and each of which exhibited a minimum of four mismatches with a low probability score. Founder animals were identified by sequencing PCR product from genomic DNA. Two independent founder mice generated on a C57BL/6J background (Jackson Laboratory) were identified and crossed to WT C57BL/6J mice to test germline transmission. Four F1 heterozygotes were found to be positive for the knock-in allele. These heterozygotes were mated to derive distinct F2 populations that were subsequently intercrossed to obtain litters that were homozygous for the knock-in allele. Subsequent crosses of homozygous knock-in mice yielded healthy offspring.
Suggested Control Mice
C57BL/6J or wild type siblings
MMRRC Genetic QC Summary
The MMRRC Centers have developed a genetic QC pipeline using MiniMUGA array genotyping to provide additional information on strain backgrounds for MMRRC congenic and inbred strains. For more information on when data may be available, or to request genotyping for a strain of interest, please contact mmrrc@ucdavis.edu. Older strains may not have this information.
  • Immunology and Inflammation
Donor
Stefanie Vogel, Ph.D., University of Maryland School of Medicine.
Primary Reference

Richard K, Piepenbrink KH, Shirey KA, Gopalakrishnan A, Nallar S, Prantner DJ, Perkins DJ, Lai W, Vlk A, Toshchakov VY, Feng C, Fanaroff R, Medvedev AE, Blanco JCG, Vogel SN. A mouse model of human TLR4 D299G/T399I SNPs reveals mechanisms of altered LPS and pathogen responses. J Exp Med. 2021 Feb 1;218(2):e20200675. doi: 10.1084/jem.20200675. (Medline PMCID: 7685774)

Colony and Husbandry Information

Homozygous trio mating, wean at 4 weeks due to smaller pup size.

Colony Surveillance Program and Current Health Reports

Mice recovered from a cryo-archive will have health surveillance performed on recipient females. Health reports will be provided prior to shipment. If you require additional health status information, please email mmrrc@ucdavis.edu.
Coat Color
Black
Eye
Black
MMRRC Breeding System
Random intra-strain mating
Overall Breeding Performance
Good
Viability and Fertility: Female Male Comments
Homozygotes are viable: Yes Yes
Homozygotes are fertile: Yes Yes
Heterozygotes are fertile: Yes Yes
Age Reproductive Decline: 10 to 12 months 10 to 12 months
Average litter size
7 to 9
Recommended wean age
4 Weeks
Average Pups Weaned
7 to 9

Order Request Information

Limited quantities of breeder mice (recovered litter) are available from a cryoarchive; recovered litter usually available to ship in 3 to 4 months.

Cryopreserved material may be available upon request, please inquire to mmrrc@ucdavis.edu for more information.

A Commercial License Agreement from the Donor is required for for-profit entities to use this strain. For more information, please contact Mark Wozniak.

A Commercial License Agreement from the Donor is required for for-profit entities to use this strain. For more information, please contact Mark Wozniak

Distribution of this strain requires submission of the MMRRC Conditions of Use (COU). A link to the COU web form will be provided via email after an order has been placed; the form should be completed then or the email forwarded to your institutional official for completion.

Additional charges may apply for any special requests. Shipping costs are in addition to the basic distribution/resuscitation fees. Information on shipping costs and any additional charges will be provided by the supplying MMRRC facility.

Click button to Request this one strain. (Use the MMRRC Catalog Search to request more than one strain.)
MMRRC Item # Description Distribution Fee / Unit (US $)
*Shipping & Handling not included*
Units Notes
071385-UCD-SPERM Cryo-preserved spermatozoa $546.25 / $869.88
Non-Profit / For-Profit
Aliquot Approximate quantity3
071385-UCD-RESUS Litter recovered from cryo-archive $4,044.00 / $7,650.23
Non-Profit / For-Profit
Litter Recovered litter4; additional fees for any special requests.
Cryopreserved material may be available upon request, please inquire to mmrrc@ucdavis.edu for more information.

1 The distribution fee covers the expense of rederiving mice from a live mouse; you will receive the resulting litter. The litter will contain at minimum one mutant carrier; the actual number of animals and the gender and genotype ratios will vary. (Typically, multiple breeder pairs can be established from the recovered litter.) Prior to shipment, the MMRRC will provide information about the animals recovered. If you anticipate or find that you need to request specific genotypes, genders or quantities of mice in excess of what is likely from a resuscitated litter, you may discuss available options and pricing with the supplying MMRRC facility.

2 An aliquot contains a sufficient number of embryos (in one or more vials or straws and based on the transfer success rate of the MMRRC facility) to transfer into one to three recipients. The MMRRC makes no guarantee concerning embryo transfer success experienced in the recipient investigator's laboratory. Neither gender nor genotype ratios are guaranteed.

3 An aliquot is one straw or vial with sufficient sperm to recover at least one litter of mice, as per provided protocols, when performed at the MMRRC facility. The MMRRC makes no guarantee concerning the success of these procedures when performed outside the MMRRC facilities.

4 The distribution fee covers the expense of resuscitating mice from the cryo-archive; you will receive the resulting litter. The litter will contain at minimum one mutant carrier; the actual number of animals and the gender and genotype ratios will vary. (Typically, multiple breeder pairs can be established from the recovered litter.) Prior to shipment, the MMRRC will provide information about the animals recovered. If you anticipate or find that you need to request specific genotypes, genders or quantities of mice in excess of what is likely from a resuscitated litter, you may discuss available options and pricing with the supplying MMRRC facility.

To request material from the MMRRC: Please fill out our on-line request form (accessible from the catalog search results page, or click the Request this Strain button in the fees section). If you have questions or need assistance completing this form, you may call Customer Service at (800) 910-2291 (in USA or Canada) or (530) 757-5710 (international calls). Before you call, please have with you: the MMRRC item number, quantity needed, Bill-to and Ship-to contact information.