R26FlpoER mice express a tamoxifen-inducible, optimized FLPe recombinase variant (Flpo). Following tamoxifen administration, efficient FLP-mediated recombination is observed in all embryonic and adult tissues.
IMR Colony, The Jackson Laboratory
Dr. Alexandra L Joyner, Memorial Sloan-Kettering Cancer Center
Genetic Background | Generation |
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Allele Type | Gene Symbol | Gene Name |
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Targeted (Recombinase-expressing, Inducible) | Gt(ROSA)26Sor | gene trap ROSA 26, Philippe Soriano |
Stock No. 019016 is a C57BL/6NJ-congenic R26FlpoER strain. The phenotype for R26FlpoER mice on a mixed genetic background (mostly Swiss Webster; Stock No. 018906) is described below. In an attempt to offer alleles on well-characterized or multiple genetic backgrounds, alleles are frequently moved to a genetic background different from that on which an allele was first characterized. It should be noted that the phenotype of C57BL/6NJ-congenic R26FlpoER mice could vary from that originally described for R26FlpoER mice on a mixed genetic background. We may modify the strain description if necessary as published results become available.
Heterozygous R26FlpoER mice are viable and fertile with no reported phenotypic abnormalities. Homozygous R26FlpoER mice are viable and fertile with no expected phenotypic abnormalities (the phenotype of homozygotes has not been fully characterized to date [September 2012]). The R26FlpoER allele is designed with the CMV enhancer/chicken beta-actin core promoter and tamoxifen-inducible FlpoER fusion protein inserted into the Gt(ROSA)26Sor locus. The FlpoER fusion protein is an optimized FLPe variant (Flpo) fused to the G400V/M543A/L544A triple mutation of the human estrogen receptor ligand binding domain (ERT2) by a linker sequence that promotes FLP recombination efficiency. Under control of the endogenous Gt(ROSA)26Sor promoter/enhancer regions and the CMV enhancer/chicken beta-actin core promoter, widespread expression of FlpoER is observed in all embryonic and adult tissues, but translocation to the nucleus / FLP-mediated recombination is rare prior to tamoxifen treatment. Following tamoxifen administration, FLP-mediated recombination is observed in all embryonic and adult tissues. Importantly, recombination is consistent between animals within each stage and organ (and the extent of recombination is specific to each organ and stage of induction). When R26FlpoER mice are bred with mice containing frt-flanked sequences, tamoxifen-inducible FLP-mediated recombination will result in deletion of frt-flanked sequences in the FLP-expressing cells of the offspring.
These mice are useful for "Mosaic mutant Analysis with Spatial and Temporal control of Recombination" (MASTR). By breeding these ubiquitous inducible R26FlpoER mice with FLP-inducible eGFPcre mice (R26MASTR; Stock No. 018903), the double mutant line can be bred to mice harboring a floxed gene of interest for mosaic mutant analysis of deleted genes in any tissue at any time (embryonic or adult). For example, mice harboring the R26FlpoER, R26MASTR, and the Smolox conditional knockout allele (Stock No. 004526) exhibit sonic hedgehog receptor ablation following tamoxifen administration, and allow one to test the cell autonomous function of Smo in granule cell precursor proliferation.
The FlpoER fusion protein consists of the mouse codon-optimized FLPe recombinase variant (FLPo) fused to the G400V/M543A/L544A triple mutation of the human estrogen receptor ligand binding domain (ERT2) by a linker sequence that promotes FLP recombination efficiency. The ERT2 does not bind its natural ligand (17β-estradiol) at physiological concentrations but will bind the synthetic estrogen receptor ligands 4-hydroxytamoxifen (OHT or tamoxifen) and, with lesser sensitivity, ICI 182780. Restricted to the cytoplasm, FlpoER can only gain access to the nuclear compartment after exposure to tamoxifen. To counteract the mixed estrogen agonist effects of tamoxifen injections, which can result in late fetal abortions in pregnant mice, progesterone may be coadministered.
A targeting construct was designed with the human cytomegalovirus (CMV) enhancer/chicken beta-actin core promoter (from pCA-HZ2), the FlpoER fusion protein cDNA sequence, an SV40 polyA sequence, and a loxP-flanked PGK-neo (STOP) cassette.
The FlpoER was constructed with the FLPo DNA sequence (mouse codon-optimized FLPe recombinase variant of the Saccharomyces cerevisiae FLP1 recombinase gene; modified for translation in mammalian cells and higher recombination efficiency in cells and in mice) fused to the ERT2 sequence (G400V/M543A/L544A triple mutation of the human estrogen receptor ligand binding domain from bp 2082-2090 of CreERT2) by a linker sequence (TGCGTACGCGGATCC; encoding the CVRGS peptide optimized for FLP recombination efficiency). The linker sequence joins the last Flpo codon and the first ERT2 codon. This targeting construct was inserted into the Gt(ROSA)26Sor locus via electroporation into 129S6/SvEvTac-derived CY2.3 embryonic stem (ES) cells. Correctly targeted ES cells were injected into recipient blastocysts and chimeric mice were bred with C57BL/6 mice to establish the R26FlpoER colony. The mutant mice were then bred to Swiss Webster outbred mice and/or Swiss Webster mice with other reporter alleles for many generations. In addition, the donating investigator reports that although these mice harbor no additional mutant loci, they may contain incidental genetic background contributions as a result of past matings with other mutant mouse strains.
R26FlpoER mice on a mixed genetic background (mostly Swiss Webster) were sent to The Jackson Laboratory Repository in 2012 as Stock No. 018906. Upon arrival, some R26FlpoER mice were backcrossed to C57BL/6NJ inbred mice (Stock No. 005304) for several generations to create this C57BL/6NJ-congenic R26FlpoER colony at The Jackson Laboratory Repository as Stock No. 019016. As of July 2014, nine generations of backcrossing to C57BL/6NJ had been performed.
Expressed Gene | FLP, FLP recombinase, Saccharomyces cerevisiae |
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Site of Expression | Following tamoxifen administration, FLP-mediated recombination is observed in all embryonic and adult tissues. |
Allele Name | targeted mutation 3, Alexandra L Joyner |
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Allele Type | Targeted (Recombinase-expressing, Inducible) |
Allele Synonym(s) | Gt(ROSA)26Sortm3(ACTB-FLPo/ERT2)Alj; R26FlpoER |
Gene Symbol and Name | Gt(ROSA)26Sor, gene trap ROSA 26, Philippe Soriano |
Gene Synonym(s) | |
Expressed Gene | FLP, FLP recombinase, Saccharomyces cerevisiae |
Site of Expression | Following tamoxifen administration, FLP-mediated recombination is observed in all embryonic and adult tissues. |
Strain of Origin | 129S6/SvEvTac |
Chromosome | 6 |
General Note | ES cell line = CY2.3 (129S6/SvEvTac-derived: gift of Rockefeller Gene Targeting Core) |
Molecular Note | The sequence encoding a FLPo/ERT2 fusion protein (optimized FLP recombinase fused to the tamoxifen-responsive mutated human estrogen receptor ligand-binding domain), SV40 polyadenylation site and a loxP-flanked PGK-neo cassette was cloned downstream of the human CMV enhancer/chicken ACTB promoter (CAG). This construct was targeted to the Gt(ROSA)26Sor locus in embryonic stem cells. |
Mutations Made By | Dr. Alexandra Joyner, Memorial Sloan-Kettering Cancer Center |
Mutant mice were bred to C57BL/6NJ inbred mice (Stock No. 005304) for many generations to establish this congenic strain. When maintaining the live congenic colony, heterozygous or homozygous mice may be bred together. The coat color of these mice is expected to be black.
When using the R26FlpoER mouse strain in a publication, please cite the originating article(s) and include JAX stock #019016 in your Materials and Methods section.
Facility Barrier Level Descriptions
Service/Product | Description | Price |
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Heterozygous or wildtype for Gt(ROSA)26Sor<tm3(CAG-FLPo/ERT2)Alj> |
Frozen Mouse Embryo | B6N.129S6(Cg)-Gt(ROSA)26Sor<tm3(CAG-flpo/ERT2)Alj>/J Frozen | $2595.00 |
Frozen Mouse Embryo | B6N.129S6(Cg)-Gt(ROSA)26Sor<tm3(CAG-flpo/ERT2)Alj>/J Frozen | $2595.00 |
Frozen Mouse Embryo | B6N.129S6(Cg)-Gt(ROSA)26Sor<tm3(CAG-flpo/ERT2)Alj>/J Frozen | $3373.50 |
Frozen Mouse Embryo | B6N.129S6(Cg)-Gt(ROSA)26Sor<tm3(CAG-flpo/ERT2)Alj>/J Frozen | $3373.50 |
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