C57BL/6-Myd88^tm1.1Rein/J

Stock number: 029349
Product name: Myd88^c-pL252P
Research areas: Cancer Research, Immunology, Inflammation and Autoimmunity Research, Research Tools

Description

The conditional knockin Myd88^c-pL252P allele has loxP sites flanking exons 2-6 of the Myd88 gene and a duplicated exon 2-6 containing the amino acid substitution L252P. Removal of the floxed sequence allows expression of the L252P mutation. The orthologous human mutation, L265P, is associated with B-cell malignancies, and, in particular, diffuse large B-cell lymphoma (DLBCL).

Detailed description

The knockin Myd88^c-pL252P allele conditionally expresses the L252P mutation. In humans, the orthologous mutation in Myd88, L265P, is associated with B cell malignancies. Cre-mediated excision of the loxP flanked exon 2-6 enables expression of a duplicated exon 2-6 carrying the L25P mutation. The Myd88 gene encodes a cytosolic adaptor protein that functions as a signal transducer relaying Toll-like receptor signaling to activation of NFKB. 29% of activated B cell type diffuse large B-cell lymphomas (DLBCL) carry the human L265P mutation and are characterized by constitutive activation of the NFKB pathway. Mice homozygous for the floxed allele are viable and fertile with no reported abnormalities.

When bred to mice expressing B cell specific Cre recombinase, Cd19^tm1(cre)Cgn (#006785), the resulting mice exhibit lymphoproliferative disease, splenomegaly and the occasional clonal lymphoma beginning at 60 weeks of age. Median survival is reduced to 501 days as compared to 539 days. A similar phenotype is observed when bred with mice carrying Tg(Cr2-cre)3Cgn (#006368) or Aicda^tm1(cre)Mnz alleles. These mice may be useful in studying B cell neoplastic disease.

Development

A targeting vector was designed by Dr. Christian Reinhardt (University of Cologne) to have a loxP site upstream of exon 2, followed by an FRT-flanked neo cassette. An F3-flanked puromycin cassette followed by a second loxP site is located downstream of exon 6 of the myeloid differentiation primary response gene 88 gene (Myd88). Exons 2-6 were duplicated and inserted downstream of the second loxP site, and an L252P amino acid substitution (orthologous to the human L265P mutation) was introduced into duplicated exon 5.

The construct was electroporated into C57BL/6NTac-derived embryonic stem (ES) cells. Correctly targeted ES cells were injected into recipient blastocysts, and chimeric males were bred with C57BL/6-Tg(CAG-Flpe)2Arte transgenic mice to delete the neo and puromycin cassettes.

The resulting conditional Myd88^c-pL252P mice were bred with C57BL/6 wildtype mice for multiple generations (and the Flp-expressing transgene was removed) prior to sending males to The Jackson Laboratory Repository in 2016.

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).

Allele

Symbol: Myd88^tm1.1Rein
Name: targeted mutation 1.1, H Christian Reinhardt
MGI accession ID: MGI:5776121
Generation method: Targeted
Attributes: Conditional ready (e.g. floxed); Humanized sequence; No functional change
Marker symbol: Myd88
Marker name: myeloid differentiation primary response gene 88
Chromosome: 9
Strain of origin: C57BL/6NTac
Molecular note: The targeting vector was designed to have a loxP site upstream of exon 2, followed by an FRT-flanked neo cassette. An F3-flanked puromycin cassette followed by a second loxP site is located downstream of exon 6. Exons 2-6 were duplicated and inserted downstream of the second loxP site, and an L252P amino acid substitution (orthologous to the human L265P mutation) was introduced into duplicated exon 5. Flp-mediated recombination removed the neo and puromycin cassettes.