C57BL/6-Gt(ROSA)26Sor^{tm1(CAG-ECFP*/Rapgef3/Venus*)Kama}/J

Stock number: 032205
Product name: CAMPER, cAMP Encoded Reporter
Research areas: Cell Biology Research, Neurobiology Research, Research Tools

Description

When these CAMPER, cAMP Encoded Reporter, mice are crossed with a Cre recombinase expressing strain, the floxed-STOP cassette is deleted and the Gt(ROSA)26Sor and CAG promoters drive expression of the TEpacVV cAMP sensor in the Cre-expressing cells. CAMPER mice allow selective cAMP concentration changes monitoring via FRET imaging in a tissue/cell-specific manner and may be useful in studies of G-protein-coupled receptors (GPCRs) responses.

Detailed description

Intracellular signaling second messenger Cyclic AMP (cAMP) activates protein kinase A, cyclic nucleotide regulated ion channel, and EPAC proteins, including EPAC1, also known as Rap guanine nucleotide exchange factor (GEF) 3. CAMPER, cAMP Encoded Reporter, mice serve as a cre recombinase inducible FRET indicator strain for cAMP concentration changes. These mice contain sequence encoding the ^TEpac^VV cAMP sensor inserted into the Gt(ROSA)26Sor locus. Expression of the ^TEpac^VV is blocked by a loxP-flanked STOP fragment. The ^TEpac^VV cAMP FRET sensor consists of an mTurquoise donor and ^cp173Venus-Venus acceptor sandwiching the cAMP-binding protein Rapgef3, (also known as Epac1). Homozygotes are viable and fertile.

When bred to a strain with Cre recombinase expression in medium spiny neurons or injected with AAV-Cre virus, this mutant mouse strain may be useful in studies of neuromodulation in many neuronal populations.

 

Development

A targeting vector designed containing (5’ to 3’): CAG promoter, floxed STOP cassette, ^TEpac^VV cAMP sensor, WPRE, polyadenylation signal sequence, was inserted into the GT(ROSA)26Sor locus (Ai9 Addgene plasmid # 22799). The ^TEpac^VV cAMP FRET sensor consists of an mTurquoise donor and ^cp173Venus-Venus acceptor sandwiching the cAMP-binding protein Rapgef3, (also known as Epac1).

The construct was electroporated into C57BL/6 derived C2 embryonic stem (ES) cells. Correctly targeted ES cells were injected into blastocysts. The resulting chimeric animals were tested for germline transmission. The mice were backcrossed to C57BL/6 for 2 generations. Upon arrival, sperm was cryopreserved. To establish our live colony, an aliquot of frozen sperm was used to fertilize C57BL/6NJ oocytes (Stock No. 005304).

Allele

Symbol: Gt(ROSA)26Sor^{tm1(CAG-ECFP*/Rapgef3/Venus*)Kama}
Name: targeted mutation 1, Kirill Martemyanov
MGI accession ID: MGI:6194564
Generation method: Targeted
Attributes: Conditional ready (e.g. floxed); Reporter
Synonyms: CAMPER
Marker symbol: Gt(ROSA)26Sor
Marker name: gene trap ROSA 26, Philippe Soriano
Marker synonyms: beta geo; Gtrgeo26; Gtrosa26; R26; ROSA26; SETD5-AS1; Thumpd3as1
Chromosome: 6
Strain of origin: C57BL/6NTac
Molecular note: The targeting vector contains from (5' to 3'): CAG promoter, a loxP STOP loxP cassette, Epac cAMP sensor, WPRE, polyadenylation signal sequence and is inserted between exons 1 and 2 of the Gt(ROSA)26Sor Ai9 targeting vector. The Epac sensor reports cAMP binding by changes in fluorescence energy transfer (FRET). The sensor consists of an mTurqoise donor, a monomeric variant of the cyan fluorescent protein, and a circularly permutated Venus enhanced yellow fluorescent protein, Venus-Venus, acceptor sandwiching the cAMP-binding protein Rapgef3.