C57BL/6J-Clec4f^{em1(cre)Glass}/J

Stock number: 033296
Product name: Clec4f-Cre-tdTomato
Research areas: Developmental Biology Research, Research Tools

Description

Clec4f-Cre-tdTomato knock-in mice express Cre recombinase and tdTomato-NLS under direction of the C-type lectin domain family 4, member f (Clec4f) promoter in Kupffer cells of the liver.

Detailed description

Clec4f-Cre-tdTomato knock-in mice express Cre recombinase and tdTomato-NLS (tdTomato tagged with nuclear localization signal) under direction of the Kupffer cell specific C-type lectin domain family 4, member f (Clec4f) promoter. Kupffer cells are liver resident macrophages that are involved in iron metabolism and the clearance of gut-derived microbial products. Homozygotes are viable and fertile. Fluorescence is detectable by flow cytometry. When bred with mice containing loxP-flanked sequences, cre-mediated recombination will result in deletion of the floxed sequences in the Cre-expressing cells of the offspring.

When bred to ROSA26iDTR mice (Stock No. 007900), resulting mice express the diphtheria toxin receptor (DTR) specifically in Kupffer cells. Treatment of those double mutant mice with diphtheria toxin (DT) results in nearly complete ablation of the Kupffer cell population within 12 hours. This results in rapid colonization of the empty niche by circulating monocytes and their subsequent differentiation to Kupffer-like cells. The double mutant mice are useful when looking at changes in repopulating liver macrophages following acute Kupffer cell depletion as a means to infer signaling pathways and transcription factors that promote Kupffer cell differentiation.

Development

Plasmids encoding signal guide RNAs, designed to insert an internal ribosome entry site (IRES) fused to a Cre recombinase sequence, a viral 2A oligopeptide (T2A) that mediates ribosomal skipping, and a tdTomato-NLS (nuclear localization signal tagged variant of red fluorescent protein (RFP) sequence) into the 3’ UTR of the C-type lectin domain family 4, member f (Clec4f) gene, and the cas9 nuclease were introduced into the cytoplasm of C57BL/6J-derived fertilized eggs with well recognized pronuclei. Targeted embryos were transferred to pseudopregnant females. Correctly targeted pups were identified by sequencing and PCR and further backcrossed to C57BL/6J mice to develop a congenic colony of Clec4f-Cre-tdTomato mice. Upon arrival at The Jackson Laboratory, 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: Clec4f^{em1(cre)Glass}
Name: endonuclease-mediated mutation 1, Christopher Glass
MGI accession ID: MGI:6283832
Generation method: Endonuclease-mediated
Attributes: Recombinase-expressing; Reporter
Synonyms: Clec4f-cre-tdTomato
Marker symbol: Clec4f
Marker name: C-type lectin domain family 4, member f
Marker synonyms: CLECSF13; D18063; Kclr; KCR; kupffer cell receptor
Chromosome: 6
Strain of origin: C57BL/6J
Site of expression: TdTomato and cre expression in Kupffer cells of the liver.
Molecular note: CRISPR/Cas9 genome editing is used to insert an internal ribosome entry site (IRES) fused to a Cre recombinase sequence, a viral 2A oligopeptide (T2A) that mediates ribosomal skipping, and a tdTomato variant of red fluorescent protein (RFP) sequence into the 3'UTR of the gene.