C57BL/6-Orai1^tm1Rao/J

Stock number: 023352
Product name: Orai1 KO
Research areas: Cell Biology Research, Developmental Biology Research, Research Tools

Description

This Orai1 knock-out strain is useful in studies of regulation of cytosolic and endoplasmic reticulum calcium levels, store-operated calcium channels and T cell and B cell function.

Detailed description

The targeted Orai1 gene encodes a pore subunit of the store-operated calcium channel, CRAC (calcium release-activated calcium) channel. Mutations in this gene have been associated with primary immunodeficiency-9 and tubular aggregate myopathy-2. These knock-out mice carry an allele of Orai1 in which exon 1 has been excised. No gene product (mRNA) is detected by RT-PCR analysis of CD4+ T cells and CD19+ B cells from homozygotes. Fewer than expected homozygotes on the C57BL/6 background are born, and homozygotes are perinatal lethal, with all homozygous pups dying by 1.5 days after birth. MEFs isolated from homozygotes exhibit significantly impaired store-operated Ca²⁺ influx.

Development

A targeting vector was designed to disrupt exon 1 of the targeted gene with a loxP site flanked self-deleting selection cassette (containing a neomycin resistance gene and Cre recombinase under the control of a testes-specific angiotensin-converting enzyme promoter). The construct was electroporated into C57BL/6NTac derived B6/3 embryonic stem (ES) cells. Correctly targeted ES cells were injected into blastocysts. The resulting chimeric animals were tested for germline transmission. The resulting chimeric animals were bred to C57BL/6J to achieve germline transmission and allow the excision of the self-deleting selection cassette. 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: Orai1^tm1Rao
Name: targeted mutation 1, Anjana Rao
MGI accession ID: MGI:3810487
Generation method: Targeted
Attributes: Null/Knockout
Marker symbol: Orai1
Marker name: ORAI calcium release-activated calcium modulator 1
Chromosome: 5
Strain of origin: C57BL/6NTac
Molecular note: Exon 1 was replaced with a self-deleting neomycin cassette that was excised in the germ line of male mice carrying the targeted allele. Gene inactivation was confirmed by a lack mRNA transcript in CD4⁺ T cells and B cells. No protein expression was detected in the skin by immunohistochemical analysis.
General note: ES cell line = B6/3.