STOCK Gt(ROSA)26Sor^tm1Phep/J

Stock number: 026570
Research areas: Neurobiology Research, Research Tools

Description

These Rosa26^SNAPCaaX reporter mice conditionally express membrane-localized SNAP-tag from the Gt(ROSA)26Sor promoter upon cre-mediated excision of a floxed stop cassette. Cells expressing the SNAP-tag can be selectively labeled through treatment with benzylguanine (BG)-linked compounds, including synthetic fluorophores.

Detailed description

SNAP-tag is a mutant form of O⁶-alkylguanine-DNA alkyltransferase (a DNA repair protein) that can, when fused to proteins of interest, be used for specific labeling of cells. SNAP-tag reacts specifically and rapidly with benzylguanine (BG) which can be linked with a range of synthetic labels, including fluorophores. The labels transfer from BG derivatives to the SNAP-tag in an irreversible, covalent fashion, enabling fast and quantitative one-step labeling of the protein of interest with minimal off-target signal and low cellular toxicity.

These Rosa26^SNAPCaaX cre reporter mice conditionally express SNAP-tag associated with a membrane-associated CaaX consensus sequence driven from the Gt(ROSA)26Sor promoter. Cre-mediated excision of a floxed stop cassette enables expression of membrane-associated SNAP-tag in the tissue(s) of interest and permits selective labeling upon application of a BG-linked compound.

Crosses with deleter and peripheral/central nervous-specific cre strains (e.g. NFH-cre, Avil-cre, Vglut2-cre) have demonstrated strong, sustained, cell-specific BG-fluorophore labeling capabilities. Labeling tolerates standard histological sample preparation and attains fluorescence intensities greater than those of more standard fluorescent protein reporters.

In addition for fluorophore labeling, applications may also include chromophore-assisted light activation (CALI) for ablation of cells, and the use of synthetic calcium or voltage sensors for neuronal activity imaging.

Development

A targeting construct introduced the following components, in order, to intron 1 of the Gt(ROSA)26Sor gene: CAG-FRT-loxP-stop-loxP-(SNAP-tag/CaaX)-WPRE-pA-AttB-Pr-FRT-Neo-pA-AttP. The mutation was introduced via homologous recombination in A9 mixed 129P2 and B6 background embryonic stem cells. This strain was maintained on a mixed genetic background by the donating laboratory.

Allele

Symbol: Gt(ROSA)26Sor^tm1Phep
Name: targeted mutation 1, Paul A Heppenstall
MGI accession ID: MGI:5604517
Generation method: Targeted
Attributes: Conditional ready (e.g. floxed); Reporter; No functional change
Marker symbol: Gt(ROSA)26Sor
Marker name: gene trap ROSA 26, Philippe Soriano
Chromosome: 6
Strain of origin: (C57BL/6 x 129P2/OlaHsd)F1
Expressed genes: SNAP-tag,SNAP-tag,
Molecular note: A targeting construct introduced the following components, in order, to intron 1 of the Gt(ROSA)26Sor gene: CAG-FRT-loxP-stop-loxP-(SNAP-tag/CaaX)-WPRE-pA-AttB-Pr-FRT-Neo-pA-AttP. The SNAP-Tag construct contains a membrane associated CaaX consensus sequence.