Heterozygous mice harbor an Lgr5-EGFP-IRES-creERT2 "knock-in" allele that both abolishes Lgr5 gene function and expresses EGFP and CreERT2 fusion protein. When these mice mice are bred with mice containing a loxP-flanked sequence of interest, tamoxifen-inducible, Cre-mediated recombination will result in deletion of the floxed sequences in the Lgr5-expressing cells of the offspring. These mice may be useful for lineage-tracing or marking Lgr5-expressing stem cells of the small intestine.
Hans Clevers, Hubrecht Institute
Genetic Background | Generation |
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N11+N4F5
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Allele Type | Gene Symbol | Gene Name |
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Targeted (Recombinase-expressing, Reporter, Inducible) | Lgr5 | leucine rich repeat containing G protein coupled receptor 5 |
Starting at:
$270.00 Domestic price for female 4-week |
348.51 Domestic price for breeder pair |
The donating investigator reports variegated expression of the Lgr5-EGFP-IRES-CreERT2 transgene in the small intestine and colon (something which may happen often with genes that are expressed early during intestinal development). This variegated expression is advantageous for performing clonal lineage tracing and sorting intestinal stem cells, but may have limitations for more quantitative studies such as Lgr5-Cre driven knockout strategies.
While homozygous mice are not viable, heterozygous Lgr5-EGFP-IRES-CreERT2 mice are viable and fertile; harboring a Lgr5-EGFP-IRES-creERT2 "knock-in" allele that both abolishes Lgr5 (Gpr49) gene function and expresses EGFP and CreERT2 fusion protein from the Lgr5 promoter/enhancer elements. EGFP fluorescence is observed in crypt base columnar cells in small intestine (aka stem cells of the small intestine) and colon. Cre-ERT2 fusion gene activity is inducible; observed in the same cells only following tamoxifen administration. EGFP or inducible CreERT2 expression may also be observed in other Lgr5-expressing cell types (including pre-malignant mouse adenomas, colon cancer cells, epithelial stem cells of the stomach gland, basal epithelial layer stem cells of the mammary glands, and hair follicle stem cells).
The donating investigator reports variegated expression of the Lgr5-EGFP-IRES-CreERT2 transgene in the small intestine and colon (something which may happen often with genes that are expressed early during intestinal development). This variegated expression is advantageous for performing clonal lineage tracing and sorting intestinal stem cells, but may have limitations for more quantitative studies such as Lgr5-Cre driven knockout strategies.
The Cre-ERT2 fusion protein consists of Cre recombinase fused to a triple mutant form of the human estrogen receptor; which 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, Cre-ERT2 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. As such, when Lgr5-EGFP-IRES-creERT2 mice are bred with mice containing loxP-flanked sequence of interest, tamoxifen-inducible, Cre-mediated recombination will result in deletion of the floxed sequences in the Lgr5-expressing cells of the offspring.
View cre expression characterization.
The EGFP-IRES-creERT2 targeting construct was designed to insert an enhanced green fluorescent protein sequence (EGFP), an internal ribosome entry site (IRES), a CreERT2 fusion gene (Cre-ERT2; Cre recombinase fused to a G400V/M543A/L544A triple mutation of the human estrogen receptor ligand binding domain), a polyA signal, and a loxP-flanked neo cassette into the first ATG codon of the targeted gene. This construct was inserted into the targeted gene via electroporation into male 129P2/OlaHsd-derived IB10/E14IB10 embryonic stem (ES) cells. Correctly targeted ES cells were injected into recipient blastocysts and chimeric males were bred with C57BL/6 females. Mutant mice were then crossed to EIIa-cre mice (C57BL/6 genetic background; see Stock No. 003724) to remove the neo selection cassette. The donating investigator reported that the resulting Lgr5-EGFP-IRES-creERT2 mice (floxed-neo removed) were then backcrossed to C57BL/6J (see SNP note below) for at least 4 generations. Next, Lgr5-EGFP-IRES-creERT2 mice were bred with Rosa26-lacZ mice (C57BL/6 genetic background; see Stock No. 003474). Mice with both mutations were further backcrossed for more than 6 generations. Upon arrival at The Jackson Laboratory, the mice were bred with C57BL/6J inbred mice (Stock No. 000664) for at least one generation to establish the colony. Mice were then selectively bred to remove the Rosa26-lacZ allele.
A 32 SNP (single nucleotide polymorphism) panel analysis, with 27 markers covering all 19 chromosomes and the X chromosome, as well as 5 markers that distinguish between the C57BL/6J and C57BL/6N substrains, was performed on the rederived living colony at The Jackson Laboratory Repository. While the 27 markers throughout the genome suggested a C57BL/6 genetic background, 2 of 5 markers that determine C57BL/6J from C57BL/6N were found to be segregating. These data suggest the mice sent to The Jackson Laboratory Repository were on a mixed C57BL/6J ; C57BL/6N genetic background.
Expressed Gene | GFP, Green Fluorescent Protein, |
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Expressed Gene | cre/ERT2, Cre recombinase and estrogen receptor 1 (human) fusion gene, |
Site of Expression | Variegated EGFP fluorescence is observed in crypt base columnar cells in small intestine (stem cells of the small intestine) and colon. |
Allele Name | targeted mutation 1, Hans Clevers |
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Allele Type | Targeted (Recombinase-expressing, Reporter, Inducible) |
Allele Synonym(s) | Lgr5 EGFP-IRES-creERT2; Lgr5tm1(cre/ESR1)Cle; Lgr5-cre; Lgr5CREER; Lgr5-creERT; Lgr5-CreERT2-IRES-EGFP; Lgr5-EGFP; Lgr5-EGFP-IRES-creERT2 |
Gene Symbol and Name | Lgr5, leucine rich repeat containing G protein coupled receptor 5 |
Gene Synonym(s) | |
Expressed Gene | GFP, Green Fluorescent Protein, |
Expressed Gene | cre/ERT2, Cre recombinase and estrogen receptor 1 (human) fusion gene, |
Site of Expression | Variegated EGFP fluorescence is observed in crypt base columnar cells in small intestine (stem cells of the small intestine) and colon. |
Strain of Origin | 129P2/OlaHsd |
Chromosome | 10 |
Molecular Note | An EGFP-IRES-creERT2 cassette was inserted into exon 1 of the gene along with a loxP-flanked neomycin cassette. The neomycin gene was then excised in vivo by crossing with cre-deleter strain. GFP expression is driven by the endogenous promoter. Cre expression is induced in Lgr5 expressing cells by tamoxifen treatment. |
Mutations Made By | Nick Barker, Hubrecht Institute |
When maintaining a live colony, heterozygous mice may be bred together, to wildtype siblings, or to C57BL/6J inbred mice (Stock No. 000664). Homozygous mice are not viable.
When using the Lgr5-EGFP-IRES-creERT2 mouse strain in a publication, please cite the originating article(s) and include JAX stock #008875 in your Materials and Methods section.
Service/Product | Description | Price |
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Heterozygous or wildtype for Lgr5<tm1(cre/ERT2)Cle> |
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