Overview

Also Known As: Lgr5-EGFP-IRES-creERT2

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.

Donating Investigator

Hans Clevers, Hubrecht Institute

Genetic overview

Genetic Background	Generation
	N11+N4F4 (2018-11-30 00:00:00)

Lgr5^{tm1(cre/ERT2)Cle}

Allele Type	Gene Symbol	Gene Name
Targeted (Recombinase-expressing, Reporter, Inducible)	Lgr5	leucine rich repeat containing G protein coupled receptor 5

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Research Applications

Research Tools
Cancer Research
Internal/Organ Research

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Base Price

Starting at:

$270.00 Domestic price for female 4-week

348.51 Domestic price for breeder pair

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Details

Important Note

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.

Detailed Description

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.

Development

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-ER^T2; 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.

Expression Data

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.

Control Suggestions

Wild-type from the colony
000664 C57BL/6J

Additional Information

Considerations for Choosing Controls

Selected References

Barker N; van Es JH; Kuipers J; Kujala P; van den Born M; Cozijnsen M; Haegebarth A; Korving J; Begthel H; Peters PJ; Clevers H. 2007. Identification of stem cells in small intestine and colon by marker gene Lgr5. Nature 449(7165):1003-7PubMed: 17934449 MGI: J:127123

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Genetics

Lgr5^{tm1(cre/ERT2)Cle}

Allele Symbol: Lgr5^{tm1(cre/ERT2)Cle}

Allele Name	targeted mutation 1, Hans Clevers
Allele Type	Targeted (Recombinase-expressing, Reporter, Inducible)
Allele Synonym(s)	targeted mutation 1, Hans Clevers; Lgr5^{tm1(cre/ERT2)Cle}
Gene Symbol and Name	Lgr5, leucine rich repeat containing G protein coupled receptor 5
Gene Synonym(s)	Gpr49; GPR49; GPR67; HG38; GRP49; FEX; G protein-coupled receptor 49; Gpr49
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

Disease/Phenotype

Disease Terms

Research Areas By Genotype

This mouse can be used to support research in many areas including:

Research Tools
- Cre-lox System
  - Cre Recombinase Expression
  - Cre Recombinase Expression: Inducible
- Genetics Research
  - Mutagenesis and Transgenesis: Cre-lox System
  - Tissue/Cell Markers: Cre-lox System
  - Tissue/Cell Markers: transplantation marker for embryonic and adult tissue
  - Mutagenesis and Transgenesis
  - Tissue/Cell Markers
  - Tissue/Cell Markers: multiple
- Developmental Biology Research
  - transplantation marker for embryonic and adult tissue
  - Cre-lox System
- Toxicology Research
  - xenograft/transplant host
- Cancer Research
- Fluorescent Proteins
Cancer Research
- Other
  - tumor metastasis
Internal/Organ Research
- Gastrointestinal Defects

Mammalian Phenotype Terms by Genotype

References

Barker N; van Es JH; Kuipers J; Kujala P; van den Born M; Cozijnsen M; Haegebarth A; Korving J; Begthel H; Peters PJ; Clevers H. 2007. Identification of stem cells in small intestine and colon by marker gene Lgr5. Nature 449(7165):1003-7PubMed: 17934449 MGI: J:127123

Additional References

Zheng Y; Song Y; Han Q; Liu W; Xu J; Yu Z; Zhang R; Li N. 2018. Intestinal epithelial cell-specific IGF1 promotes the expansion of intestinal stem cells during epithelial regeneration and functions on the intestinal immune homeostasis. Am J Physiol Endocrinol Metab 315(4):E638-E649PubMed: 29783855 MGI: J:265953
Zhao B; Chen Y; Jiang N; Yang L; Sun S; Zhang Y; Wen Z; Ray L; Liu H; Hou G; Lin X. 2019. Znhit1 controls intestinal stem cell maintenance by regulating H2A.Z incorporation. Nat Commun 10(1):1071PubMed: 30842416 MGI: J:276456
Cuitino MC; Pecot T; Sun D; Kladney R; Okano-Uchida T; Shinde N; Saeed R; Perez-Castro AJ; Webb A; Liu T; Bae SI; Clijsters L; Selner N; Coppola V; Timmers C; Ostrowski MC; Pagano M; Leone G. 2019. Two Distinct E2F Transcriptional Modules Drive Cell Cycles and Differentiation. Cell Rep 27(12):3547-3560.e5PubMed: 31130414 MGI: J:276975
Goto N; Fukuda A; Yamaga Y; Yoshikawa T; Maruno T; Maekawa H; Inamoto S; Kawada K; Sakai Y; Miyoshi H; Taketo MM; Chiba T; Seno H. 2019. Lineage tracing and targeting of IL17RB(+) tuft cell-like human colorectal cancer stem cells. Proc Natl Acad Sci U S A 116(26):12996-13005PubMed: 31182574 MGI: J:277412

Additional - Lgr5^{tm1(cre/ERT2)Cle} related

Technical Support

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Genotyping Protocols
- MELT: Lgr5^{tm1(cre/ERT2)Cle} alternate2
- Genotyping resources and troubleshooting
Dietary Information
- LabDiet® 5K52 formulation (6% fat)
Breeding Considerations

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.
- Additional Breeding and Husbandry Support
Mating System
- +/+ sibling x Heterozygote
Citation
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.

Animal Health Reports

Facility Barrier Level Descriptions

AX10 (Standard)

Pricing & Availability

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Cryorecovery - Pricing

Service	Genotype	Price
	Heterozygous or wildtype for Lgr5<tm1(cre/ERT2)Cle>

We will fulfill your order by providing at least two carriers for each strain ordered. The total number, sex, and genotypes provided will vary, although typically 8 or more animals are provided. Please check genotypes which will be recovered. While the genotypes of all animals produced will be communicated to you prior to scheduling shipment, the genotypes of animals provided may not reflect the mating scheme and genotypes described in the strain description. Animals are typically ready to ship in 11-14 weeks. If a second recovery is required to produce the minimum number of animals, then delivery time would increase to approximately 25 weeks. If we fail to produce animals of the correct genotype, you will not be charged. We cannot guarantee the reproductive success of mice shipped to your facility. If the mice are lost after the first three days (post-arrival) or do not produce progeny at your facility, a new order and fee will be necessary.

Cryorecovery to establish a Dedicated Supply for greater quantities of mice. Mice recovered can be used to establish a dedicated colony to contractually supply you mice according to your requirements. Price by quotation.

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Also Known As: Lgr5-EGFP-IRES-creERT2

Donating Investigator

Genetic overview

Research Applications

Base Price

Important Note

Detailed Description

Development

Expression Data

Control Suggestions

Additional Information

Selected References

Lgr5tm1(cre/ERT2)Cle

Allele Symbol: Lgr5tm1(cre/ERT2)Cle

Disease Terms

Research Areas By Genotype

This mouse can be used to support research in many areas including:

Mammalian Phenotype Terms by Genotype

References

Additional References

Additional - Lgr5tm1(cre/ERT2)Cle related

Genotyping Protocols

Dietary Information

Breeding Considerations

Mating System

Citation

Animal Health Reports

Live Mouse

Breeder Pair

Cryorecovery - Pricing

Payment Terms and Conditions

The Jackson Laboratory's Genotype Promise

Terms of Use

Additional Use Restrictions Apply

Licensing Information

JAX® Mice, Products & Services Conditions of Use

No Warranty

Credit for PRODUCTS or SERVICES

Animal Care and Use for SERVICES

No Liability

Lgr5^{tm1(cre/ERT2)Cle}

Allele Symbol: Lgr5^{tm1(cre/ERT2)Cle}

Additional - Lgr5^{tm1(cre/ERT2)Cle} related