C57BL/6J-Gzma^em1Asuh/J

Stock number: 038375
Product name: Gzma^S211A
Strain synonyms: C57BL/6J-GzmaS211A
Research areas: Apoptosis Research, Cell Biology Research, Immunology, Inflammation and Autoimmunity Research

Description

Gzma^S211A knock-in mice contain a CRISPR/Cas9 generated insertion of the S211A point mutation in granzyme A (Gzma) gene, resulting in a loss of GZMA enzymatic function. This strain may be useful for studying cell-mediated cytotoxicity.

Detailed description

Cytotoxic T-cells mediate apoptosis of virus or tumor infected cells via several independent pathways, one of which is a perforin-mediated pathway. In this apoptotic pathway, the pore-forming protein perforin acts together with proteases, Granzyme A and B, which eventually lead to target cell death. Granzyme A, (GZMA), coded by Gzma, activates caspase-independent programmed cell death, that begins in the mitochondria. GZMA also has proinflammatory activity, resulting in expression of pro-interleukin-1β. These Gzma^S211A knock-in (KI) mice contain a CRISPR/Cas9 generated amino acid substitution of a serine to alanine at residue 211 (S211A) in the (Gzma) gene; this mutation does not disrupt GZMA expression but results in a loss of GZMA enzymatic function.

Homozygous Gzma^S211A KI mice are viable and fertile. These mice may be useful for studying the proteolytic activity of GZMA in programmed cell-death processes.

Development

The Gzma^S211A knock-in (KI) allele was generated using CRISPR/Cas9 endonuclease-mediated genome editing. A guide RNA (GAGGGCTGCCGGCGTCCCC) was used to target the granzyme A (Gzma) locus on chromosome 13, resulting in an amino acid substitution of a serine to alanine at residue 211 (S211A). ENSMUSG00000023132 was used as reference for the exon number and guide sequences. The sgRNAs, ssDNA and the Cas9 nuclease mRNA were introduced into the cytoplasm of C57BL/6J (Stock No. 000664) zygotes with well recognized pronuclei. Correctly targeted pups were identified by PCR and Sanger sequencing. The donating laboratory backcrossed the Gzma^S211A KI mice to C57BL/6J for at least 1 generation to reduce off-target events. Upon arrival at The Jackson Laboratory, sperm was cryopreserved. To establish our live colony, an aliquot of frozen sperm was used to fertilize C57BL/6J oocytes.

Allele

Symbol: Gzma^em1Asuh
Name: endonuclease-mediated mutation 1, Andreas Suhrbier
MGI accession ID: MGI:6885562
Generation method: Endonuclease-mediated
Attributes: Not Specified
Synonyms: Gzma^S211A
Marker symbol: Gzma
Marker name: granzyme A
Marker synonyms: BLT esterase; CTLA3; Ctla-3; H factor; Hanukah factor; Hf; HFSP; SE1; serine esterase 1; TSP1; TSP-1
Chromosome: 13
Strain of origin: C57BL/6J
Molecular note: CRISPR-targeting introduced the mutation that produces the amino acid substitution of serine to alanine at position 211 (S211A).