B6;129S-Gt(ROSA)26Sor^{tm1.1(CAG-COX8A/Dendra2)Dcc}/J

Stock number: 018397
Product name: PhaM excised
Research areas: Research Tools

Description

PhAM^excised (photo-activatable mitochondria) mice express a mitochondrial-specific version of Dendra2 green/red photoswitchable monomeric fluorescent protein in mitochondria. Exposure of mitochondria to 405 nm laser light irreversibly switches green fluorescence to red, enabling measurements of mitochondrial fusion and transport.

Detailed description

Dendra2 is a green-to-red, irreversibly photoswitchable monomeric fluorescent protein derived from octocoral Dendronephthya sp. Its fusion with the mitochondrial targeting signal of subunit VIII of cytochrome c oxidase generates "mito-Dendra2", capable of mitochondrial-specific fluorescence.

In this PhAM^excised strain, mito-Dendra2 is introduced into the ubiquitously-expressed Gt(ROSA)26Sor gene downstream of a CAG (CMV-βactin) promoter. All organs isolated from these animals (e.g. central nervous system, skeletal muscle, heart, testis, lung, kidney, thymus) exhibit bright green fluorescence localized specifically to the mitochondrial compartment. Homozygotes are viable and fertile.

When a sub-population of mitochondria is exposed to 405 nm laser light, the green signal photo-converts to red fluorescence, enabling measurements of mitochondrial fusion (indicated by mixing of green and red signals) and transport.

Development

Dendra2 (a green to red irreversibly photoswitchable monomeric fluorescent protein derived from octocoral Dendronephthya sp.) fused to the mitochondrial targeting signal of subunit VIII of cytochrome c oxidase (mito-Dendra2), bovine growth hormone (bGH) polyadenylation signal, and an FRT-flanked neomycin cassette were introduced to Gt(ROSA)26Sor downstream of a CAG (CMV-βactin) promoter and loxP-flanked termination signal. The mutation was created in TC1 129S6/SvEvTac-derived embryonic stem cells. Crosses with a C57BL/6 background FLP strain excised the FRT-flanked neomycin cassette. Crosses with a germline cre deleter strain (Meox2-cre) excised the remaining floxed termination cassette. This strain was maintained on a mixed C57BL/6-129 genetic background by the donating lab.

Allele

Symbol: Gt(ROSA)26Sor^{tm1.1(CAG-COX8A/Dendra2)Dcc}
Name: targeted mutation 1.1, David C Chan
MGI accession ID: MGI:5318241
Generation method: Targeted
Attributes: Reporter
Marker symbol: Gt(ROSA)26Sor
Marker name: gene trap ROSA 26, Philippe Soriano
Chromosome: 6
Strain of origin: 129S6/SvEvTac
Expressed genes: Dendra,Dendra photoswitchable fluorescent protein,octocoral Dendronephthya
Site of expression: Cre excision of the stop signal results in expression of Dendra2, a green-to-red irreversibly photo-switchable fluorescent protein, in the mitochondria of cre-expressing tissues.
Molecular note: Dendra2 (a green to red irreversibly photoswitchable monomeric fluorescent protein derived from octocoral Dendronephthya sp.) fused to the mitochondrial targeting signal of subunit VIII of cytochrome c oxidase (mito-Dendra2), bovine growth hormone (bGH) polyadenylation signal, and an FRT-flanked neomycin cassette were introduced to Gt(ROSA)26Sor downstream of a CAG (CMV-beta-actin) promoter and loxP-flanked termination signal. Flp-mediated recombination removed the neo cassette and cre-mediated recombination excised the floxed termination cassette.