B6;129S-Gt(ROSA)26Sor^tm1.1Ksvo/J

Stock number: 023139
Product name: Rosa26 LSL H2B mCherry
Research areas: Neurobiology Research, Research Tools

Description

R26 LSL H2B mCherry (Rosa26-CAG-LSL-H2B-mCherry) mutant mice have a loxP-flanked STOP sequence followed by an H2B-mCherry fusion protein inserted into the Gt(ROSA)26Sor locus. Cre inducible, nuclear expression of the mCherry protein allows the monitoring of Cre activity in living tissues and makes possible lineage tracing of such cells in embryos, young and adult mice at desired time points.

While designed to have the floxed-STOP sequence preventing widespread expression of mCherry in the absence of Cre recombinase, low levels of mCherry expression are detectable prior to introduction of Cre recombinase. Importantly, after exposure to Cre recombinase, the mCherry expression levels in the nucleus have been shown to be significantly greater than those baseline levels (e.g., when bred to the strong germline Cre expressing Sox2-Cre mouse line [Stock No. 008454]). As such, it is recommended that researchers include Cre-negative R26 LSL H2B mCherry controls to establish the baseline mCherry levels in their experiments.

Detailed description

R26 LSL H2B mCherry was designed with a CAG promoter and loxP-flanked STOP sequence upstream of a nuclear-targeted H2B-mCherry fusion protein, all inserted into the Gt(ROSA)26Sor locus. While designed to have the floxed-STOP sequence preventing widespread expression of mCherry in the absence of Cre recombinase, low levels of mCherry expression are detectable prior to introduction of Cre recombinase. Importantly, after exposure to Cre recombinase, the mCherry expression levels in the nucleus have been shown to be significantly greater than those baseline levels (e.g., when bred to the strong germline Cre expressing Sox2-Cre mouse line [Stock No. 008454]). As such, it is recommended that researchers include Cre-negative R26 LSL H2B mCherry controls to establish the baseline mCherry levels in their experiments.
Mice that are homozygous for the R26 LSL H2B mCherry mutant allele are viable and fertile. These R26 LSL H2B mCherry mice may be useful in monitoring the Cre expression in living tissues, and tracing the lineage of such cells in embryos, young, and adult mice at desired time points.

Development

A targeting vector was designed to contain an H2B-mCherry fusion protein (coding sequence for the human HIST1H2BB gene [histone 1 H2bb] followed C-terminally by Enhanced Red Fluorescent Protein gene) downstream of a CAG promoter and loxP-flanked stop sequence (neomycin resistance gene and a trimer of the SV40 polyadenylation sequence). An AttB-PGKNeo-AttP cassette was placed downstream of the H2B-mCherry fusion protein. This entire construct was inserted into the Gt(ROSA)26Sor locus via electroporation of 129S6-derived embryonic stem (ES) cells. Correctly targeted ES cells were microinjected in blastocysts and chimeric males were bred to a C57BL/6 females. Offspring were bred to B6.129S4-Gt(ROSA)26Sor^{tm3(phiC31*)Sor}/J mice (Stock No. 007743) to remove the neo cassette. Resulting mice were bred to C57BL/6J mice and then were crossed to remove the phiC31-expressing mutation. Upon arrival at The Jackson Laboratory, these mice were bred to C57BL/6J mice (Stock No. 000664) for at least one generation to establish a colony.

Allele

Symbol: Gt(ROSA)26Sor^tm1.1Ksvo
Name: targeted mutation 1.1, Karel Svoboda
MGI accession ID: MGI:5512720
Generation method: Targeted
Attributes: Conditional ready (e.g. floxed); Reporter; Humanized sequence; No functional change
Marker symbol: Gt(ROSA)26Sor
Marker name: gene trap ROSA 26, Philippe Soriano
Chromosome: 6
Strain of origin: 129S6/SvEvTac
Expressed genes: RFP,Red Fluorescent Protein,coral; H2BC3,H2B clustered histone 3,human
Molecular note: A targeting vector was designed to contain a CAG promoter, followed by a loxP-STOP-LoxP (LSL) sequence and an H2B-mCherry fusion protein (coding sequence for the human HIST1H2BB gene [histone 1 H2bb] followed C-terminally by Enhanced Red Fluorescent Protein gene) An AttB-PGKNeo-AttP cassette was placed downstream of the H2B-mCherry fusion protein. This entire construct was inserted into the Gt(ROSA)26Sor locus. PhiC31-mediated recombination removed the neomycin resistance cassette.