B6.Cg-Il21^tm1.1Hm/DcrJ

Stock number: 030295
Product name: B6.IL21-VFP knock-in reporter
Research areas: Immunology, Inflammation and Autoimmunity Research, Research Tools

Description

This IL21-VFP knock-in strain expresses VFP (vivid Verde fluorescent protein) from the endogenous Il21 locus and is useful as a reporter for IL21-expressing CD4 T follicular helper (T_FH) cells, which are the precursors of mature T_FH cells.

Detailed description

The targeted Il21 gene encodes an immunoregulatory cytokine important in innate and adaptive immune responses. Mutations in this gene have been associated with common variable immunodeficiency-11. These IL21-VFP mice express VFP, vivid Verde fluorescent protein, from the endogenous Il21 locus. Green fluorescence is detected by flow cytometry in a large number of CD44+ICOS+CD4+ T cells and approximately 40% of PD1+ cells in immunologically challenged IL21-VFP mice. When crossed to an autoimmune disease model strain, immunohistochemical analysis of spleen from the double mutant mice shows that the VFP fluorescence co-localizes with IL21 expressing cells. Heterozygotes and homozygotes exhibit similar percentages of VFP positive cells. Homozygotes are viable and fertile. Expression of the endogenous Il21 gene is not altered.

Green-emitting VFP was isolated from the tropical scleractinian coral Cyphastrea microphthalma, rapidly matures at 37°C and exhibits a maximum excitation peak at 491nm and a maximum emission peak at 503 nm, with brighter fluorescence than EGFP.

Development

A targeting vector containing an IRES sequence, VFP (vivid Verde fluorescent protein) sequence, a loxP site flanked PGK-neo cassette, was inserted into the non-coding region of exon 5 of the targeted gene. The construct was electroporated into unspecified C57BL6/N-derived embryonic stem (ES) cells. Correctly targeted ES cells were injected into C57BL/6 blastocysts. The resulting chimeric animals were tested for germline transmission. The mice were crossed to germ-line expressing Cre recombinase transgenic mice, on the C57BL/6 genetic background, to excise the selection cassette. The mice were crossed to C57BL/6 to breed away from the Cre recombinase transgene. Heterozygotes were intercrossed to generate homozygotes.

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). SNP (single nucleotide polymorphism) analysis performed on the rederived living colony at The Jackson Laboratory revealed that the strain is on a C57BL/6J background.

Allele

Symbol: Il21^tm1.1Hm
Name: targeted mutation 1.1, Herbert Morse III
MGI accession ID: MGI:6095616
Generation method: Targeted
Attributes: Reporter
Synonyms: IL21-VFP
Marker symbol: Il21
Marker name: interleukin 21
Marker synonyms: CVID11; IL-21; Za11
Chromosome: 3
Strain of origin: B6.Cg-Thy1^a
Expressed genes: GFP,Green Fluorescent Protein,
Site of expression: Green fluorescence is detected by flow cytometry in a large number of CD44+ICOS+CD4+ T cells and approximately 40% of PD1+ cells in immunologically challenged Il21^tm1.1Hm mice.
Molecular note: The targeting vector contains an IRES sequence, VFP (vivid Verde fluorescent protein) sequence, and a loxP-flanked PGK-neo cassette inserted into the non-coding region of exon 5. Cre-mediated recombination removed the floxed neo cassette.