B6J.B6NTac(SJL)-Srsf2^tm1.1Oaw/J

Stock number: 028376
Product name: Srsf2^P95H floxed
Research areas: Cancer Research, Cell Biology Research, Research Tools

Description

Srsf2^P95H floxed mice possess loxP sites flanking the coding region of the Srsf2 gene, and also contains a downstream mutant coding region. Removal of the floxed wild type exons allows expression of the mutant sequence. This strain may be useful for studying myelodysplastic syndromes.

Detailed description

Srsf2^P95H floxed mice possess loxP sites flanking the endogenous coding region of the serine/arginine-rich splicing factor 2 (Srsf2) gene. They also contain a duplication of the coding region containing a proline to histidine mutation downstream of the 3’ loxP site. SRSF2 encodes a spliceosomal gene associated with the regulation of consititive splicing. Mutations in SRSF2 have been associated with the onset and pathogenesis of myelodysplastic syndromes (MDS) and chronic myelomonocytic leukemia (CMML). Srsf2^P95H floxed mice that are heterozygous for this allele are viable and fertile, the donating investigator has not attempted to make homozygotes. When bred to mice that express tissue-specific Cre recombinase, resulting offspring will have endogenous exons deleted, and will instead express the mutant Srsf2^P95H gene in the cre-expressing tissues.

When bred to B6.Cg-Tg(Mx1-cre)1Cgn/J mice (from Stock No. 003556), expressing Cre recombinase in hematopoietic stem cells (HSCs) after administration of poly(IC), mice develop a mild anemia with macrocytosis and leukopenia, as well as morpholgic dysplasia as seen in human myelodysplastic syndromes.

Development

A targeting vector was designed by Dr. Omar Abdel-Wahab (Memorial Sloan Kettering Cancer Center) to insert a loxP site upstream of exon 1 of the serine/arginine-rich splicing factor 2 (Srsf2) gene. cDNA encoding the sequence of exon 3, and an SV40 polyadenylation sequence bound to a frt-flanked neomycin resistance (neo) cassette, followed by a second loxP site were inserted downstream of exon 2. Duplicate exons 1-2 were inserted downstream of the neo cassette. This second exon 1 contains a point mutation in amino acid 95 which alters the nucleotide sequence from CCG to CAC, resulting in an amino acid change from Proline to Histidine. The cDNA for exon 3 was fused in frame to endogenous exon 2. The construct was electroporated into C57BL/6NTac-derived IC1 embryonic stem (ES) cells. Correctly targeted ES cells were injected into Balb/c blastocysts and resulting chimeric mice were bred with B6.Cg-Tg(ACTFLPe)9205Dym/J transgenic mice (Stock No. 005703) to delete the neo cassette. Resulting progeny were crossed to remove the Flp-expressing transgene, and Srsf2^P95H floxed offspring were backcrossed to C57BL/6 mice for at least 6 generations (see SNP note below). 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).

A 32 SNP (single nucleotide polymorphism) panel analysis, with 27 markers covering all 19 chromosomes and the X chromosome, as well as 5 markers that distinguish between the C57BL/6J and C57BL/6N substrains, was performed on the rederived living colony at The Jackson Laboratory Repository. While the 27 markers throughout the genome suggested a C57BL/6 genetic background, 4 of 5 markers that determine C57BL/6J from C57BL/6N were found to be segregating. These data suggest the mice sent to The Jackson Laboratory Repository were on a mixed C57BL/6J ; C57BL/6N genetic background.

Allele

Symbol: Srsf2^tm1.1Oaw
Name: targeted mutation 1.1, Omar Abdel-Wahab
MGI accession ID: MGI:5694699
Generation method: Targeted
Attributes: Conditional ready (e.g. floxed)
Marker symbol: Srsf2
Marker name: serine/arginine-rich splicing factor 2
Chromosome: 11
Strain of origin: C57BL/6NTac
Molecular note: A targeting vector was designed to insert a loxP site upstream of exon 1, cDNA encoding the sequence of exon 3 fused in frame to the endogenous exon 2, followed by an SV40 polyadenylation sequence bound to an FRT-flanked neomycin resistance (neo) cassette and a second loxP site. Duplicate exons 1-2 were inserted downstream of the second loxP site and upstream of the endogenous exon 3. The second exon 1 contains a mutation in amino acid 95 which alters the nucleotide sequence from CCG to CAC, resulting in an amino acid change from Proline to Histidine. Flp-mediated recombination removed the FRT-flanked neo cassette.