These TRPM8EGFPf targeted mutation (knock in) mice display deficits in cool thermosensation and may be useful in studies of cold nociception.
Ardem Pataoputian, The Scripps Research Institute
Genetic Background | Generation |
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|
Allele Type | Gene Symbol | Gene Name |
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Targeted (Reporter, Null/Knockout) | Trpm8 | transient receptor potential cation channel, subfamily M, member 8 |
These mutant mice express farnesylated enhanced GFP gene (EGFPf) from the endogenous Trpm8 locus. Mice that are homozygous for the targeted mutation are viable, fertile, normal in size and do not display any gross physical or behavioral abnormalities. No wildtype gene product (mRNA) is detected although low levels of a truncated mRNA splice variant is sometimes detected by RT-PCR analysis of total RNA derived from dorsal root ganglia (DRG) isolated from homozygotes. No Trpm8 locus mRNA is detected by in situ hybridization of DRG isolated from homozygotes. In thermotaxis assays performed in a cage equipped with surface temperature gradients, homozygous mice spend less time in the warmer area compared to wildtype controls. Homozygotes display diminished responses to application of acetone and injection of icilin. Noxious cold sensation of colder temperatures (sub zero) is normal. Mutant mice do not display cold-induced analgesia in response to pain. EGFPf-positive neurons isolated from homozygotes are more sensitive to cold and menthol. TRPM8-expressing neurons isolated from mutants are specifically labeled by EGFPf expression. EGFPf fluorescence is detected in the dorsal root ganglia and spinal cord from both homozygotes and heterozygotes (with an increased fluorescence intensity observed in homozygotes). Axonal terminals are visualized utilizing immunohistochemical methods.
A targeting vector containing a farnesylated enhanced GFP gene (EGFP-F), SV40polyA signal sequence, and a loxP site flanked EM7-PGK-Neo cassette, was used to replace sequence encoding amino acids 2-29. The construct was electroporated into 129S1/Sv-Oca2+ Tyr+ Kitl+ derived CJ7 embryonic stem (ES) cells. Correctly targeted ES cells were injected into C57BL/6 blastocysts. The resulting male chimeric animals were crossed to C57BL/6 female mice. The mice were crossed to an EIIA-Cre deleter strain (Stock No. 003724) to remove the floxed Neo cassette. Upon arrival at The Jackson Laboratory, the mice were crossed to C57BL/6J (Stock No. 000664) at least once to establish the colony.
Expressed Gene | GFP, Green Fluorescent Protein, |
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Site of Expression | Farnesylated EGFP (EGFPf) fluorescence is detected in the dorsal root ganglia and spinal cord. |
Allele Name | targeted mutation 1, Ardem Patapoutian |
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Allele Type | Targeted (Reporter, Null/Knockout) |
Allele Synonym(s) | Trpm8-; Trpm8EGFPf |
Gene Symbol and Name | Trpm8, transient receptor potential cation channel, subfamily M, member 8 |
Gene Synonym(s) | |
Expressed Gene | GFP, Green Fluorescent Protein, |
Site of Expression | Farnesylated EGFP (EGFPf) fluorescence is detected in the dorsal root ganglia and spinal cord. |
Strain of Origin | 129S1/Sv-Oca2+ Tyr+ Kitl+ |
Chromosome | 1 |
Molecular Note | Velocigene technology was used to delete sequences encoding amino acid residues 2-29 and insert a farnesylated enhanced GFP gene followed by an SV40polyA tail in frame with the start codon of the endogenous gene. RT-PCR confirmed absence of wild-type transcript in mutants, however, in some samples a truncated splice variant was detected at low levels. |
Mutations Made By | Ardem Patapoutian, The Scripps Research Institute |
When maintaining a live colony, these mice can be bred as homozygotes.
When using the STOCK Trpm8tm1Apat/J mouse strain in a publication, please cite the originating article(s) and include JAX stock #014581 in your Materials and Methods section.
Facility Barrier Level Descriptions
Service/Product | Description | Price |
---|---|---|
Heterozygous or Wild-type for Trpm8<tm1Apat> |
Frozen Mouse Embryo | STOCK Trpm8<tm1Apat>/J Frozen Embryo | $2595.00 |
Frozen Mouse Embryo | STOCK Trpm8<tm1Apat>/J Frozen Embryo | $2595.00 |
Frozen Mouse Embryo | STOCK Trpm8<tm1Apat>/J Frozen Embryo | $3373.50 |
Frozen Mouse Embryo | STOCK Trpm8<tm1Apat>/J Frozen Embryo | $3373.50 |
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