B6(129S4)-Fcgr1^{em1.1(icre)Moli}/J

Stock number: 039826
Product name: Fcgr1-cre
Research areas: Cancer Research, Research Tools

Description

Fcgr1-cre knock-in mice have an IRES-tdTomato-T2A-iCre inserted at the C-terminal of the Fcgr1 gene. They are useful for deletion of floxed sequences in macrophages and for fluorescent tracing of Fcgr1 expression in studies of tumor-associated macrophage differentiation.

Detailed description

Fcgr1-cre knock-in mice have an IRES-tdTomato-T2A-iCre (internal ribosome entry site, tandem dimer Tomato Red Fluorescent Protein, T2A self-cleaving peptide and a codon-improved iCre recombinase gene) inserted at the C-terminal of the Fc receptor, IgG, high affinity I (Fcgr1) gene. The insertion does not affect endogenous gene expression. The Fcgr1, also known as CD64, is a high-affinity immunoglobulin gamma Fc receptor expressed constitutively in monocytes and macrophages. Mice heterozygous and homozygotes for the Fcgr1-cre allele are viable and fertile with no overt phenotype. The donating laboratory reported that tdTomato can be detected by flow cytometry in Fcgr1-expressing tissue-resident macrophages. These mice can be used to fluorescently mark macrophage nuclei and when bred with mice containing loxP-flanked sequence, iCre-mediated recombination will result in the deletion of the floxed sequences in macrophages.

Development

Fcgr1-cre mice were generated by CRISPR/Cas9-mediated homologous recombination. The targeting construct contained IRES, tdTomato, T2A self-cleaving peptide, iCre recombinase and an FRT-flanked PGK-neo-BGHpA cassette. This was designed to insert downstream of the stop codon of the Fcgr1 gene. Double guide RNAs targeting at sites around the stop codon (5'-TTGTGCCCCAGTACTGTCAC-3') and (5'-CAAAGTTGACCCTGAAACTG-3') were cloned into the pX335 vector for a double nickase-targeting strategy. Mouse Fcgr1-201 transcript (ENSMUST00000029748.8) was used as a reference for designing the guide sequences. The targeting construct and pX335 vectors were co-injected into albino C57BL/6 embryonic stem (ES) cells. ES cell clones with proper insertion at 108 bp downstream of the Fcgr1 stop codon and stable chromosomes were injected into pseudo-pregnant recipient female mice mixed with non-albino C57BL/6 ES cells. Successful offspring chimera harboring the targeted allele were genotype confirmed and mated to wildtype C57BL/6 mice. Offspring were subsequently crossed with Rosa26-FLPe mice (Stock No. 003946) to remove the PGK-neo-BGHpA cassette. See The Jackson Laboratory quality control note below. The donating laboratory reports maintaining the Fcgr1-cre mice on a C57BL/6 genetic background prior to sending males to The Jackson Laboratory in 2025. 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).

In 2025, quality control screening was performed on the mice sent to The Jackson Laboratory, our first generation rederived living colony and their subsequent offspring after breeding to C57BL/6J. This revealed that all heterozygous Fcgr1-cre mice were positive for a neomycin resistance (neo) sequence while all their wildtype littermates were negative for neo. Furthermore, heterozygous mice analyzed via sequencing at The Jackson Laboratory showed a single FRT sequence was present between the iCre and endogenous Fcgr1 downstream region. Taken all together, this suggests the Fcgr1-cre allele had its FRT-flanked neo removed, but retains a neo sequence closely linked to (or elsewhere within) the Fcgr1-cre allele. It has not been determined if the retained neo is a full or partial sequence, nor is it known where it lies near/within the Fcgr1-cre allele.

Allele

Symbol: Fcgr1^{em1.1(icre)Moli}
Name: endonuclease-mediated mutation 1, Ming O Li
MGI accession ID: MGI:7516071
Generation method: Endonuclease-mediated
Attributes: Recombinase-expressing; Reporter
Synonyms: Fcgr1^{em1.1(cre)Moli}; Fcgr1-cre; Cd64-iCre-tdtomato
Marker symbol: Fcgr1
Marker name: Fc receptor, IgG, high affinity I
Marker synonyms: CD64; CD64A; CD64b; FCG1; FcgammaRI; FcgammaRIa; FCGR1; Fcgr1b; FcRI; IGFR1; IGFRB
Chromosome: 3
Strain of origin: C57BL/6
Molecular note: The targeting construct inserted IRES, tdTomato, T2A self-cleaving peptide, iCre recombinase, and an FRT site-flanked PGK-NEO-BGHpA cassette downstream of the stop codon of the Fcgr1 gene by CRISPR–Cas9-mediated homologous recombination in mouse ES cells. Mice harboring the targeted allele were subsequently crossed with Rosa26FLPe mice to remove the neo cassette. However, analysis of the mice revealed that a neo sequence closely linked to (or elsewhere within) the Fcgr1-cre allele is present. It has not been determined if the retained neo is a full or partial sequence, nor is it known where it lies near/within the Fcgr1-cre allele.