NOD.Cg-Nfkbid^em3Dvs Foxp3^tm2Tch Tg(Nfkbid)60-2Dvs/DvsJ

Stock number: 041269
Product name: NOD-60A.Foxp3-GFP
Research areas: Diabetes and Obesity Research, Research Tools

Description

NOD-60A.Foxp3-GFP mice carry a knock-out mutation of the endogenous NOD Nfkbid allele in combination with a C57BL/J-derived Nfkbid transgenic allele. An additional X-linked Foxp3 GFP targeted knock-in enables fluorescent labeling of regulatory T cells (Tregs). Mice incorporating the transgene are highly resistant to diabetes. 

Detailed description

NOD-60A.Foxp3-GFP mice carry a CRISPR/cas9-generated knock-out mutation of the endogenous NOD Nfkbid (nuclear factor of kappa light polypeptide gene enhancer in B cells inhibitor, delta) allele in combination with a C57BL/J-derived Nfkbid transgenic allele. An additional X-linked Foxp3 (forkhead box P3) targeted knock-in enables the fluorescent labeling of regulatory T cells (Tregs).

The Tg(Nfkbid)60-2Dvs (60A) transgene is carried in a hemizygous state. Transgenic mice from this line are highly resistant to the diabetes associated with the parental NOD strain, have elevated levels of regulatory T cells in the periphery, and present with splenomegaly. Nontransgenic littermates lacking NFKBID protein exhibit accelerated diabetes by 20 weeks of age and have reduced levels of regulatory T cells.

It is recommended that transgenic females be paired with nontransgenic males to maintain a colony due to the aggressive diabetes phenotype in nontransgenic mice. Typically 1-2 litters are produced from breeding pairs that involve Nfkbid transgene carrier females. Freund’s adjuvant may be used if needed, e.g. if the colony is small or breeding nontransgenic females becomes necessary.

An NOD-60A nonfluorescent version of this strain, lacking the Foxp3 allele, is available as Stock No. 041268.

Development

Nfkbid^em3Dvs is a CRISPR/Cas9-generated mutation created and maintained on a NOD/ShiLtDvs genetic background that ablates functional Nfkbid mRNA expression (NOD.NfkbidKO.2A; Stock No. 041273). Nfkbid^em3Dvs is an 8-nt deletion and an insertion of 154-nt, removing the translation start site in exon 3.

To create the Tg(Nfkbid)60-2Dvs transgene, a genomic region encompassing the C57BL/6J-derived Nfkbid allele was isolated from a bacterial artificial chromosome (BAC) and cloned into a pUC57 simple plasmid containing chicken HS4 insulator and retrieval sequences. A 303-bp intergenic segment downstream of Nfkbid was removed for genotyping purposes. Nfkbid transgenic mice were generated by pronuclear injection of the excised C57BL/6J segment into NOD/ShiLtDvs embryos. Founder mice were then crossed with mice carrying the NOD.NfkbidKO.2A mutation to create compound mutant NOD-60A mice. The Nfkbid transgene is maintained in a hemizygous state. The copy number of the transgenic element appears to be 6, but the insertion site is unknown (November 2025).

NOD-60A mice were further crossed with Foxp3^EGFP mice (Stock No. 025097) created by Dr. Talal Chatila (Boston Children's Hospital) to produce these NOD-60A.Foxp3-GFP compound mutant mice. To create the Foxp3^EGFP allele, an IRES-EGFP-SV40 poly A sequence was inserted immediately downstream of the endogenous Foxp3 translational stop codon, but upstream of the endogenous polyA signal, generating a bicistronic locus encoding both Foxp3 and EGFP.

Embryos were cryopreserved (NOD/ShiLtDvs ~N8F12).

Allele

Symbol: Nfkbid^em3Dvs
Name: endonuclease-mediated mutation 3, David V Serreze
MGI accession ID: MGI:7580893
Generation method: Endonuclease-mediated
Attributes: Null/Knockout
Marker symbol: Nfkbid
Marker name: nuclear factor of kappa light polypeptide gene enhancer in B cells inhibitor, delta
Marker synonyms: IkappaBNS; IkBNS; RGD1308055; TA-NFKBH
Chromosome: 7
Strain of origin: NOD
Molecular note: CRISPR/cas9 mediated recombination using a single guide RNA (5'-AGGCCCATTTCCCCTGGTGA-3') created an 8-nt deletion and an inverted insertion of 154 nucleotides copied from exon 2 removing the translation start site in exon 3. Western blotting of thymus from homozygotes found no NFKBID protein.

Allele

Symbol: Foxp3^tm2Tch
Name: targeted mutation 2, Talal A Chatila
MGI accession ID: MGI:3699400
Generation method: Targeted
Attributes: Reporter
Synonyms: FoxP3.GFP; Foxp3^EGFP; Foxp3^IRES-GFP; Foxp3^tm2(EGFP)Tch; Foxp3-GFP KI
Marker symbol: Foxp3
Marker name: forkhead box P3
Marker synonyms: AIID; DIETER; IPEX; JM2; PIDX; RGD1562112; scurfin; XPID
Chromosome: X
Strain of origin: 129X1/SvJ
Expressed genes: GFP,Green Fluorescent Protein,
Site of expression: EGFP is expressed in the Foxp3⁺ T cell population. Some EGFP expression is noted in a small population of CD8⁺ thymocytes.
Molecular note: An IRES-EGFP-SV40 poly A sequence was inserted immediately downstream of the endogenous Foxp3 translational stop codon, but upstream of the endogenous polyA signal, generating a bicistronic locus encoding both Foxp3 and EGFP. Expression of EGFP is restricted to the T cell lineage, primarily to the CD4+ T cell population.

Allele

Symbol: Tg(Nfkbid)60-2Dvs
Name: transgene insertion 60-2, David V Serreze
MGI accession ID: MGI:8266304
Generation method: Transgenic
Attributes: Inserted expressed sequence
Marker symbol: Tg(Nfkbid)60-2Dvs
Marker name: transgene insertion 60-2, David V Serreze
Chromosome: UN
Strain of origin: NOD/ShiLtDvs
Molecular note: A genomic region encompassing the C57BL/6J-derived Nfkbid allele was isolated from a bacterial artificial chromosome (BAC) and cloned into a pUC57 simple plasmid containing chicken HS4 insulator and retrieval sequences. A 303-bp intergenic segment downstream of Nfkbid was removed for genotyping purposes.