B6.Cg-Clec9a^tm5.1Crs/J

Stock number: 037805
Product name: Clec9a^tdTomato
Research areas: Cell Biology Research, Immunology, Inflammation and Autoimmunity Research, Research Tools

Description

Clec9a^tdTomato knock-in/knock-out mice have tdTomato with a BGH_pA inserted into the ATG in exon 1 of the Clec9a gene. This strain is useful for dendritic cell lineage fate mapping and for studying adaptive immune responses to antigens in apoptotic and necrotic cells.

Detailed description

Clec9a^tdTomato knock-in/knock-out mice have tdTomato with a BGH_pA inserted into the ATG in exon 1 of the C-type lectin domain family 9, member a (Clec9a) gene, abolishing endogenous gene expression. Clec9a encodes for DNGR-1, an endocytic receptor of the C-type lectin superfamily that is selectively expressed at high levels by CD8α+ dendritic cells (DC) and at lower levels by plasmacytoid DCs. DNGR-1 recognizes an intracellular ligand that is exposed in dead cells and facilitates cross-presentation of dead-cell-associated antigens through a mechanism that involves interaction of its intracellular tail with the tyrosine kinase, SYK. Homozygous mice are viable and fertile.

In these mice, tdTomato is evident in conventional dendritic cell (cDC) subtype 1 (cDC1), which are DNGR-1+ and are nearly 100% tdTomato+, whereas cDC2, which do not express DNGR-1, are <1% labeled. Analysis of E11.5 DNGR-1 reporter embryos show rare DNGR-1+ cells embedded in the ventricular zone, which displays concurrent expression of the progenitor marker SOX2 but not the neuroepithelial marker nestin. In contrast, no DNGR-1+ cells are found in the ependymal layer of adult Clec9a^tdTomato reporter mice, indicating that expression is confined to embryonic stages. Fluorescence is detectable by immunohistochemical staining and FACS analysis.

Of note, other CLEC9A expressing lines are available:
Clec9a^gfp knock-in/knock-out mice (Stock No. 017696) have an EGFP sequence replacing the endogenous Clec9a sequence.
Clec9a^cre knock-in/knock-out mice (Stock No. 025523) have a codon improved cre recombinase sequence replacing the endogenous Clec9a sequence.
Clec9a^Cre/ERT2 knock-in (Stock No. 037804) have cre/ERT2 inserted into the C-terminus of the endogenous Clec9a sequence and retain endogenous gene expression.

Development

A targeting vector was designed to insert tdTomato with a BGH_pA into the ATG in exon 1 of the C-type lectin domain family 9, member a (Clec9a) gene. A frt-flanked neomycin resistance (neo) cassette was inserted downstream. The construct was electroporated into B6.Cg-Thy1^a-derived Bruce4 embryonic stem (ES) cells. Correctly targeted ES cells were injected into blastocysts and resulting chimeric mice were bred with FLPe-expressing mice on a C57BL/6 genetic background to remove the neo cassette. Progeny were crossed to remove the FLPe-expressing transgene and resulting Clec9a^tdTomato mice were bred to C57BL/6J mice for at least 12 generations by the donating laboratory. 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: Clec9a^tm5.1Crs
Name: targeted mutation 5.1, Caetano Reis e Sousa
MGI accession ID: MGI:7338966
Generation method: Targeted
Attributes: Reporter
Synonyms: Clec9a^tdTomato
Marker symbol: Clec9a
Marker name: C-type lectin domain family 9, member a
Marker synonyms: 9830005G06Rik; CD370; DNGR1; DNGR-1; RGD1562513; UNQ9341
Chromosome: 6
Strain of origin: B6.Cg-Thy1^a
Molecular note: The allele was generated by conventional gene targeting (knock-in) adding tdTomato with a BGH_pA at the ATG in exon 1 of the Clec9a gene. An FRT-flanked Neo cassette in the original constructs was removed by crossing to an FLPe expressing strain.