C.129S4(B6)-Ifng^tm3.1Lky/J

Stock number: 017580
Product name: Great
Research areas: Cancer Research, Cell Biology Research, Immunology, Inflammation and Autoimmunity Research, Research Tools

Description

The "interferon-gamma reporter with endogenous polyA transcript" (GREAT) allele has an IRES-eYFP reporter cassette inserted between the translational stop codon and 3' UTR/polyA tail of the interferon gamma (Ifng) gene. Thus, the bicistronic IFNγ-IRES-eYFP mRNA is under control of the endogenous IFNγ promoter/enhancer regions with proper regulation defined by the endogenous 3' UTR and polyA tail. These GREAT mice are a fluorescent reporter strain for studying innate immunity/adaptive immunity against viral and intracellular bacterial infections, for studying anti-tumor biology, as well for studying aberrant IFN-γ expression associated with autoinflammatory/autoimmune diseases.

Detailed description

The "interferon-gamma reporter with endogenous polyA transcript" (GREAT) allele has an IRES-eYFP reporter cassette inserted between the translational stop codon and 3' UTR/polyA tail of the interferon gamma (Ifng) gene. Thus, the bicistronic IFNγ-IRES-eYFP mRNA is under control of the endogenous IFNγ promoter/enhancer regions with proper regulation defined by the endogenous 3' UTR and polyA tail. Expression of eYFP is observed by FACS/immunofluorescence microscopy in Ifng-expressing cell types; including innate immune responding cells (natural killer [NK] cells, natural killer T [NKT] cells) and cytotoxic T lymphocyte (CTL) effector cells of the adaptive immune response (CD4⁺ and CD8⁺T cells).

Of note, the donating investigator reports that these GREAT mice are a more useful tool than their IFNγ-IRES-YFP-BGHpolyA knockin (YETI) mice. Because the YETI allele has a bovine growth hormone polyA tail (rather than the endogenous IFN-γ polyA tail), homozygous YETI mice exhibit aberrant bicistronic IFNγ-IRES-YFP mRNA regulation that results in high IFN-γ blood levels and premature death. In contrast, homozygous GREAT mice are viable, fertile and normal in size, with proper transcript regulation defined by the endogenous IFN-γ 3' UTR/polyA tail.

Development

A targeting vector was designed with a loxP-flanked neomycin resistance cassette, an internal ribosome entry site (IRES), and an enhanced yellow fluorescent protein (eYFP) sequence. This vector was inserted downstream of the endogenous translational stop codon and upstream of the 3' untranslated region (UTR) and polyA tail of the interferon gamma (Ifng) gene via electroporation into 129S4/SvJae-derived PrmCre embryonic stem (ES) cells. These PrmCre (or PC3) ES cells contain the Tg(Prm-cre)70Og transgene with the protamine promoter directing cre expression to the male germline. Correctly targeted ES cells were injected into recipient blastocysts. Chimeric males were bred to C57BL/6 females. The male offspring from this initial breeding lack the floxed neo cassette in their germline, and were bred to female C57BL/6 mice. Offspring with the IRES-eYFP cassette and without the Pmc-Cre transgene were identified to establish the colony of "interferon-gamma reporter with endogenous polyA transcript" (GREAT) mice. These GREAT mice were subsequently backcrossed to BALB/cJ mice for at least ten generations. Upon arrival at The Jackson Laboratory Repository, mice were bred to BALB/cJ inbred mice (Stock No. 000651) for at least one generation.

Allele

Symbol: Ifng^tm3.1Lky
Name: targeted mutation 3.1, Richard M Locksley
MGI accession ID: MGI:5317415
Generation method: Targeted
Attributes: Reporter; No functional change
Marker symbol: Ifng
Marker name: interferon gamma
Chromosome: 10
Strain of origin: 129S4/SvJae-Tg(Prm-cre)70Og
Expressed genes: YFP,Yellow Fluorescent Protein,jellyfish
Site of expression: Cells activated to express Ifng will express EYFP.
Molecular note: A targeting vector was designed with a loxP-flanked neomycin resistance cassette, an internal ribosome entry site (IRES), and an enhanced yellow fluorescent protein (eYFP) sequence. This vector was inserted downstream of the endogenous translational stop codon and upstream of the 3' untranslated region (UTR). Cre-mediated recombination removed the neo cassette.