B6(Cg)-Sting1^em1.1Ciphe/J

Stock number: 041165
Product name: STING-OST^fl
Research areas: Cancer Research, Cell Biology Research, Immunology, Inflammation and Autoimmunity Research, Research Tools

Description

STING-OST^fl mice may be useful for studying T cells that recognize tumor antigens for mounting antitumor immune responses.

Detailed description

STING-OST^fl mice have loxP sites flanking exon 3 of the stimulator of interferon response cGAMP interactor 1 (Sting1) gene, also known as the transmembrane protein 173 (Tmem173) gene. The allele also carries a Twin-Strep-tag-coding sequence (OST) appended at the 5' end of the exon 3, the first coding exon. STING protein acts as a facilitator of innate immune signaling and interferon beta (IFNβ) production through the detection of tumor antigens for mounting antitumor immune responses. Mice that are homozygous for this allele are viable and fertile. When bred to mice that express tissue-specific Cre recombinase, the resulting offspring will have exon 3 removed in the Cre-expressing tissues. Without recombination, the allele expresses a functional STING protein with a fused OST affinity tag at the N-terminus for purification and identification. The donating investigators noted that the OST-STING protein is less active than the WT STING protein in response to stimulation by cGAS or cGAMP, due to the presence of the OST tag at the N-terminus slightly interfering with the signaling capacity of the protein. These STING-OST^fl mice may be useful for studying T cells that recognize tumor antigens for mounting antitumor immune responses.

Development

The mouse Sting1 gene (also called Tmem173; ENSMUSG00000024349) was edited using a double-stranded HDR template (targeting vector) containing 867 and 1260 bp-long 5' and 3' homology arms, respectively. The STING-OST^fl targeting vector was designed to insert a loxP site and a frt-flanked neomycin resistance (neo) cassette inserted into intron 2, 110 bp upstream of the start codon, a Twin-Strep-tag-coding sequence (OST) that was appended at the 5' end of the first coding exon (exon 3), and a loxP site located in intron 3, 40 bp downstream of the 3' end of exon 3 (reference transcript: ENSMUST00000115728.5). Two sgRNA-containing pX330 plasmids (pSpCas9) with protospacer-adjacent motifs (PAM) corresponding to each sgRNA and present in the targeting vector were destroyed via silent mutations to prevent CRISPR-Cas9 cleavage. Two sgRNA-containing pX330 plasmids and a targeting vector were electroporated into C57BL/6N-derived JM8.F6 embryonic stem (ES) cells. ES cell clones with proper homologous recombination, screened by Southern blot and PCR analysis, were injected into recipient blastocysts, and resulting chimeric males were bred with C57BL/6N females. Resulting offspring were bred with mice with Flp recombinase (B6;SJL-Tg(ACTFLPe)9205Dym/J Stock No. 003800) to remove the neo cassette, and progeny were subsequently bred together to remove the FLPe-expressing transgene. Offspring were bred to C57BL/6N mice for at least five generations be the donating laboratory (see SNP note below). Upon arrival at The Jackson Laboratory Repository, sperm was cryopreserved. To establish our live colony, an aliquot of frozen sperm was used to fertilize C57BL/6NJ oocytes (Stock No. 005304).

In 2026, 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 and C57BL/6N substrains, was performed on the males sent to The Jackson Laboratory Repository. Three of the 5 markers that determine C57BL/6J from C57BL/6N were found to be segregating. These data suggest the mice sent to The Jackson Laboratory Repository were on a mixed C57BL/6N;C57BL/6J genetic background.

Allele

Symbol: Sting1^em1.1Ciphe
Name: targeted mutation 1.1, Centre d'ImmunoPhenomique
MGI accession ID: MGI:7720866
Generation method: Endonuclease-mediated
Attributes: Conditional ready (e.g. floxed); Epitope tag
Synonyms: Sting1^tm1.1Ciphe; STING-OST^fl; Tmem173^tm1Ciphe
Marker symbol: Sting1
Marker name: stimulator of interferon response cGAMP interactor 1
Marker synonyms: 2610307O08Rik; ERIS; hMITA; hSTING; MITA; MPYS; NET23; RGD1562552; rSTING; SAVI; Sting; STING-beta; Tmem173
Chromosome: 18
Strain of origin: C57BL/6N
Molecular note: A loxP site and an FRT-flanked neomycin resistance cassette were inserted in intron 2, 110 bp upstream of the start codon, a Twin-Strep-tag coding sequence (OST) was appended at the 5’ end of the first coding exon (exon 3), and a loxP site was inserted in intron 3, 40 bp downstream of the 3’ end of exon 3. Flp-mediated recombination removed the neomycin resistance cassette, leaving exon 3 floxed. The fusion of the OST to the protein can be used in affinity purification.