Overview

Also Known As: R26^rtTA;Col1a1^3F2A

Following doxycycline (dox) administration, these single-gene transgenic R26^rtTA;Col1a1^3F2A mice express the dox-inducible polycistronic 3F2A cassette (three mouse reprogramming genes Oct4 [Pou5f1], Klf4, and c-Myc [Myc]) from the Col1a1 locus. While somatic expression of these three reprogramming factors alone does not allow the generation of induced pluripotent stem cells (iPSCs), addition of other reprogramming factors (such as Sox2) allows multiple somatic cell types to be directly reprogrammed to generate iPSCs by culture in dox. As such, somatic cells derived from R26^rtTA;Col1a1^3F2A mice are useful to screen for small molecules that can replace Sox2 for reprogramming.

Donating Investigator

Rudolf Jaenisch, Whitehead Institute, Massachusetts Institute of Technology

Genetic overview

Genetic Background	Generation

Col1a1^{tm5(tetO-Pou5f1,-Klf4,-Myc)Jae}

Allele Type	Gene Symbol	Gene Name
Targeted (Inducible, Inserted expressed sequence, RMCE-ready)	Col1a1	collagen, type I, alpha 1

Gt(ROSA)26Sor^{tm1(rtTAM2)Jae}*

Allele Type	Gene Symbol	Gene Name
Targeted (Transactivator)	Gt(ROSA)26Sor	gene trap ROSA 26, Philippe Soriano

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Research Applications

Research Tools
Cancer Research
Cell Biology Research

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Base Price

Starting at:

$2,854.50 Domestic price Cryo Recovery

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Details

Detailed Description

Mice heterozygous for both targeted mutations (R26-rtTA and Col1a1::tetO-3F2A) are viable and fertile (mice homozygous for both targeted mutations were not generated by the donating investigator but are expected to be viable and fertile as well). These double mutant R26^rtTA;Col1a1^3F2A mice have widespread expression of the optimized form of reverse tetracycline-controlled transactivator (rtTA-M2) protein directed to multiple tissues by the Gt(ROSA)26Sor promoter. In the absence of the tetracycline analog doxycycline (dox), expression of the dox-inducible 3F2A cassette from the Col1a1 locus is not detected. Following dox administration, this single-gene transgenic mouse strain expresses the polycistronic 3F2A cassette (three mouse reprogramming genes Oct4 [Pou5f1], Klf4, and c-Myc [Myc]). While somatic expression of these three reprogramming factors alone does not allow the generation of induced pluripotent stem cells (iPSCs), addition of other reprogramming factors (such as Sox2) allows multiple somatic cell types to be directly reprogrammed to generate iPSCs by culture in doxycycline. As such, somatic cells derived from R26^rtTA;Col1a1^3F2A mice are useful to screen for small molecules that can replace Sox2 for reprogramming. Because the reprogramming factors are carried on a single polycistronic construct, these double mutant R26^rtTA;Col1a1^3F2A mice can be easily maintained, and the transgene can be easily transferred into other genetic backgrounds if needed.

Development

The R26-M2rtTA targeted mutation has an optimized form of reverse tetracycline controlled transactivator (rtTA-M2) followed by a β-globin intron, polyA signal and PGK-puromycin cassette, all inserted downstream of the Gt(ROSA)26Sor promoter. The targeting vector was electroporated into (C57BL/6 x 129S4Sv/Jae)F1-derived V6.5 embryonic stem (ES) cells. Next, these targeted ES cells (termed V19 ES cells) were retargeted at the 3' UTR of the Col1a1 locus for insertion of (from 5' to 3') a loxP-flanked pgkNEOpolyA cassette, the tetracycline responsive element (TRE or tetO), the single polycistronic expression cassette 3F2A, and SV40 intron plus polyA signal. The 3F2A cassette contains the three mouse reprogramming genes Oct4 (Pou5f1; POU domain, class 5, transcription factor 1), Klf4 (Kruppel-like factor 4 (gut)), and c-Myc (Myc; myelocytomatosis oncogene) separated by two different viral 2A oligopeptides that mediate ribosomal skipping (T2A [from insect Thosea asigna virus] and E2A [equine rhinitis A virus], respectively). The resulting ES cells, now targeted with rtTA-M2 in the Gt(ROSA)26Sor locus and tetO-3F2A in the Col1a1 locus, were injected into B6D2F1 tetraploid blastocysts. The double mutant chimeric males were bred with B6D2F1 females to generate the colony. Double mutant pups on the resulting mixed genetic background were sent to The Jackson Laboratory Repository. Upon arrival, double mutant R26^rtTA;Col1a1^3F2A mice were bred with B6129SF1/J mice (Stock No. 101043) for at least one generation to establish the colony. Double mutant mice were then bred together to maintain the colony.

Expression Data

Expressed Gene	Myc, myelocytomatosis oncogene, mouse, laboratory
Expressed Gene	Pou5f1, POU domain, class 5, transcription factor 1, mouse, laboratory
Expressed Gene	Klf4, Kruppel-like factor 4 (gut), mouse, laboratory
Site of Expression	widely expressed.
Expressed Gene	rtTA, reverse tetracycline-controlled transactivator, E. coli
Site of Expression	Expresses an optimized rtTA protein (rtTA-M2). Inducible target gene expression is detected in liver, bone marrow, stomach, intestine, and skin, with lower levels in the heart, lungs, kidney, spleen, and thymus; no expression is detected in the brain and testes.

Control Suggestions

Approximate Controls

Additional Information

Considerations for Choosing Controls

Selected References

Carey BW; Markoulaki S; Beard C; Hanna J; Jaenisch R. 2010. Single-gene transgenic mouse strains for reprogramming adult somatic cells. Nat Methods 7(1):56-9PubMed: 20010831 MGI: J:157298
Hochedlinger K; Yamada Y; Beard C; Jaenisch R. 2005. Ectopic expression of Oct-4 blocks progenitor-cell differentiation and causes dysplasia in epithelial tissues. Cell 121(3):465-77PubMed: 15882627 MGI: J:98920

View All References

Genetics

Col1a1^{tm5(tetO-Pou5f1,-Klf4,-Myc)Jae}

Allele Symbol: Col1a1^{tm5(tetO-Pou5f1,-Klf4,-Myc)Jae}

Allele Name	targeted mutation 5, Rudolf Jaenisch
Allele Type	Targeted (Inducible, Inserted expressed sequence, RMCE-ready)
Allele Synonym(s)	Col1a1^{tm5(tetO-Pou5f1,-Klf4,-Myc)Jae}; targeted mutation 5, Rudolf Jaenisch
Gene Symbol and Name	Col1a1, collagen, type I, alpha 1
Gene Synonym(s)	Cola1; Cola1; COLIA1; Mov-13; Col1a-1; Cola-1; Mov-13; Moloney leukemia virus 13; OI4; Col1a-1; Cola-1; OI3; EDSC; OI1; OI2; EDSARTH1
Promoter	tetO, tet operator,
Expressed Gene	Myc, myelocytomatosis oncogene, mouse, laboratory
Expressed Gene	Pou5f1, POU domain, class 5, transcription factor 1, mouse, laboratory
Expressed Gene	Klf4, Kruppel-like factor 4 (gut), mouse, laboratory
Site of Expression	widely expressed.
Strain of Origin	(C57BL/6 x 129S4/SvJae)F1
Chromosome	11
General Note	The allele was made in KH2 cells that carry Gt(ROSA)26Sor^{tm1(rtTAM2)Jae} and Col1a1^{tm2(tetO-Pou5f1)Jae} (J:159351*).
Molecular Note	RMCE on KH2 ES cells inserted a cassette containing (from 5' to 3') a loxP-flanked pgkNEOpolyA cassette, the tetracycline responsive element (TRE or tetO), the single polycistronic expression cassette 3F2A, and SV40 intron plus polyA signal into the 3' UTR. The 3F2A cassette contains the three mouse reprogramming genes Oct4 (Pou5f1; POU domain, class 5, transcription factor 1), Klf4 (Kruppel-like factor 4 (gut)), and c-Myc (Myc; myelocytomatosis oncogene) separated by two different viral 2A oligopeptides that mediate ribosomal skipping (T2A [from insect Thosea asigna virus] and E2A [equine rhinitis A virus], respectively).

Gt(ROSA)26Sor^{tm1(rtTAM2)Jae}*

Allele Symbol: Gt(ROSA)26Sor^{tm1(rtTAM2)Jae}*

Allele Name	targeted mutation 1, Rudolf Jaenisch
Allele Type	Targeted (Transactivator)
Allele Synonym(s)	Gt(ROSA)26Sor^{tm1(rtTA*M2)Jae}; targeted mutation 1, Rudolf Jaenisch
Gene Symbol and Name	Gt(ROSA)26Sor, gene trap ROSA 26, Philippe Soriano
Gene Synonym(s)	beta geo; ROSA26; R26; Gtrgeo26; Gtrosa26; Gtrgeo26; Gtrosa26; AV258896; expressed sequence AV258896; gene trap ROSA 26; gene trap ROSA b-geo 26; SETD5-AS1; Thumpd3as1
Expressed Gene	rtTA, reverse tetracycline-controlled transactivator, E. coli
Site of Expression	Expresses an optimized rtTA protein (rtTA-M2). Inducible target gene expression is detected in liver, bone marrow, stomach, intestine, and skin, with lower levels in the heart, lungs, kidney, spleen, and thymus; no expression is detected in the brain and testes.
Strain of Origin	(C57BL/6 x 129S4/SvJae)F1
Chromosome	6
General Note	This mutation was created during the generation of mutant ES cell line KH2, which also contains a frt docking site downstream of the endogenous Col1a1 locus on Chr 11 (Col1a1). The two mutations have subsequently been bred apart.
Molecular Note	An optimized form of reverse tetracycline controlled transactivator (rtTA-M2) was inserted downstream of the Gt(ROSA)26Sor promoter and was followed by a PGK-puro selection cassette. This mutant form of rtTA termed M2 has five amino acid substitutions in the tetR moiety of tTA: S12G, E19G, A56P, D148E and H179R. This mutated form of transactivator protein has increased doxycycline sensitivity. Mice have widespread expression of the rtTA-M2 protein.
Mutations Made By	Rudolf Jaenisch, Whitehead Institute, Massachusetts Institute of Technology

Disease/Phenotype

Disease Terms

Research Areas By Genotype

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

Research Tools
- Genetics Research
  - Mutagenesis and Transgenesis: transcriptional activation
  - Mutagenesis and Transgenesis: Tetop Tet System
  - Mutagenesis and Transgenesis
- Cancer Research
  - Tetop Tet System
- Tet Expression Systems
  - tTA/rtTA Expressing Strains
  - tTA/rtTA Responsive Strains
Cancer Research
- Other
Cell Biology Research
- Transcriptional Regulation

Mammalian Phenotype Terms by Genotype

The following phenotype information is associated with a similar, but not exact match to this JAX^® Mice strain

Genotype: Col1a1^{tm5(tetO-Pou5f1,-Klf4,-Myc)Jae}/Col1a1⁺ Gt(ROSA)26Sor^{tm1(rtTAM2)Jae}/Gt(ROSA)26Sor⁺
involves: 129S4/SvJae C57BL/6

mammalian phenotype

no phenotypic analysis

References

Carey BW; Markoulaki S; Beard C; Hanna J; Jaenisch R. 2010. Single-gene transgenic mouse strains for reprogramming adult somatic cells. Nat Methods 7(1):56-9PubMed: 20010831 MGI: J:157298
Hochedlinger K; Yamada Y; Beard C; Jaenisch R. 2005. Ectopic expression of Oct-4 blocks progenitor-cell differentiation and causes dysplasia in epithelial tissues. Cell 121(3):465-77PubMed: 15882627 MGI: J:98920

Additional - Col1a1^{tm5(tetO-Pou5f1,-Klf4,-Myc)Jae} related

Additional - Gt(ROSA)26Sor^{tm1(rtTAM2)Jae}* related

Technical Support

Contact Technical Support

Genotyping Protocols
- Probe:Gt(rosa)26sor^Tm1SorProbe
- Separated MCA:Gt(ROSA)26Sor^tm1Sor
- Standard PCR:Gt(ROSA)26Sor^tm1Sor
- Genotyping resources and troubleshooting
Breeding Considerations

When maintaining a live colony, mice that are homozygous for both mutations may be bred together.
- Additional Breeding and Husbandry Support
Citation
When using the R26^rtTA;Col1a1^3F2A mouse strain in a publication, please cite the originating article(s) and include JAX stock #011013 in your Materials and Methods section.

Animal Health Reports

Facility Barrier Level Descriptions

Production of mice from cryopreserved embryos or sperm occurs in a maximum barrier room, G200

Pricing & Availability

Cryo Recovery

Domestic
International

Live Mouse

Age

Genotype

Price

weeks

Cryorecovery - Pricing

Service	Genotype	Price
	Heterozygous or wildtype for Gt(ROSA)26Sor<tm1(rtTA*M2)Jae>, Heterozygous or wildtype for Col1a1<tm5(tetO-Pou5f1,-Klf4,-Myc)Jae>

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.

Related Products and Services

Frozen Mouse Embryo	STOCK Gt(ROSA)26Sor<tm1(rtTA*M2)Jae> Col1a1<tm5(tetO-Pou5f1-	$2595.00
Frozen Mouse Embryo	STOCK Gt(ROSA)26Sor<tm1(rtTA*M2)Jae> Col1a1<tm5(tetO-Pou5f1-	$2595.00
Frozen Mouse Embryo	STOCK Gt(ROSA)26Sor<tm1(rtTA*M2)Jae> Col1a1<tm5(tetO-Pou5f1-	$3373.50
Frozen Mouse Embryo	STOCK Gt(ROSA)26Sor<tm1(rtTA*M2)Jae> Col1a1<tm5(tetO-Pou5f1-	$3373.50

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The Jackson Laboratory has rigorous genetic quality control and mutant gene genotyping programs to ensure the genetic background of JAX® Mice strains as well as the genotypes of strains with identified molecular mutations. JAX® Mice strains are only made available to researchers after meeting our standards. However, the phenotype of each strain may not be fully characterized and/or captured in the strain data sheets. Therefore, we cannot guarantee a strain's phenotype will meet all expectations. To ensure that JAX® Mice will meet the needs of individual research projects or when requesting a strain that is new to your research, we suggest ordering and performing tests on a small number of mice to determine suitability for your particular project.

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Also Known As: R26rtTA;Col1a13F2A

Donating Investigator

Genetic overview

Research Applications

Base Price

Detailed Description

Development

Expression Data

Control Suggestions

Approximate Controls

Additional Information

Selected References

Col1a1tm5(tetO-Pou5f1,-Klf4,-Myc)Jae

Allele Symbol: Col1a1tm5(tetO-Pou5f1,-Klf4,-Myc)Jae

Gt(ROSA)26Sortm1(rtTA*M2)Jae

Allele Symbol: Gt(ROSA)26Sortm1(rtTA*M2)Jae

Disease Terms

Research Areas By Genotype

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

Mammalian Phenotype Terms by Genotype

Genotype: Col1a1tm5(tetO-Pou5f1,-Klf4,-Myc)Jae/Col1a1+ Gt(ROSA)26Sortm1(rtTA*M2)Jae/Gt(ROSA)26Sor+involves: 129S4/SvJae * C57BL/6

mammalian phenotype

References

Additional - Col1a1tm5(tetO-Pou5f1,-Klf4,-Myc)Jae related

Additional - Gt(ROSA)26Sortm1(rtTA*M2)Jae related

Genotyping Protocols

Breeding Considerations

Citation

Animal Health Reports

Production of mice from cryopreserved embryos or sperm occurs in a maximum barrier room, G200

Live Mouse

Cryorecovery - Pricing

Related Products and Services

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

Also Known As: R26^rtTA;Col1a1^3F2A

Col1a1^{tm5(tetO-Pou5f1,-Klf4,-Myc)Jae}

Allele Symbol: Col1a1^{tm5(tetO-Pou5f1,-Klf4,-Myc)Jae}

Gt(ROSA)26Sor^{tm1(rtTAM2)Jae}*

Allele Symbol: Gt(ROSA)26Sor^{tm1(rtTAM2)Jae}*

Genotype: Col1a1^{tm5(tetO-Pou5f1,-Klf4,-Myc)Jae}/Col1a1⁺ Gt(ROSA)26Sor^{tm1(rtTAM2)Jae}/Gt(ROSA)26Sor⁺
involves: 129S4/SvJae C57BL/6

Additional - Col1a1^{tm5(tetO-Pou5f1,-Klf4,-Myc)Jae} related

Additional - Gt(ROSA)26Sor^{tm1(rtTAM2)Jae}* related