Overview

Also Known As:c-Kit^lox66-71 (c-Kit lox66-71)

c-Kit^lox66-71 is a Cre-recombinase conditional knockout/eGFP-reporter allele - allowing cre-inducible c-Kit deletion and concomitant labeling of the c-Kit-deficient cells with eGFP. These mice are useful for imaging of c-Kit-deficient cells in vivo as well as facilitating isolation of these cells in vitro.

Donating Investigator

Shahin Rafii, Weill Cornell Medical College

Genetic overview

Genetic Background	Generation

Kit^tm1.1Sraf

Allele Type	Gene Symbol	Gene Name
Targeted (Reporter, Null/Knockout)	Kit	KIT proto-oncogene receptor tyrosine kinase

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Research Applications

Research Tools

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Details

Detailed Description

c-Kit^lox66-71 is a Cre-recombinase conditional knockout/eGFP-reporter allele - allowing cre-inducible c-Kit deletion and concomitant labeling of the c-Kit-deficient cells with eGFP.

Prior to Cre recombination, the c-Kit^lox66-71 allele will transcribe a full-length c-Kit transcript, and no eGFP expression is reported.

Exposure to Cre recombinase will invert the fragment bordered by the inward-facing lox71 and lox66 sites (lox71-SA-IRES-eGFP-2pA-lox66) into sense orientation; creating the eGFP-expressing c-Kit knockout allele (c-Kit^ΔGFP). Specifically, the transcriptional stops upstream of the exons encoding the transmembrane and kinase domains of c-Kit result in a truncated protein and loss of c-Kit cell surface expression. The splice-acceptor-IRES sequences lead to eGFP expression in c-Kit-expressing cells (direct fluorescence). The donating investigator reports eGFP direct fluorescence is observable by flow cytometry, but weak by imaging. These mice are useful for imaging of c-Kit-deficient cells in vivo as well as facilitating isolation of these cells in vitro.

Development

The c-Kit lox66-71 allele was designed by Dr. Shahin Rafii (Weill Cornell Medical College). A targeting vector was designed to insert the following, all in reverse transcriptional orientation, into intron 8 of the kit oncogene locus (Kit; c-Kit): frt-flanked PGK-neo-pA cassette, inward-facing lox71 site, splice-acceptor, IRES-eGFP-2pA, inward-facing lox66 site. C57BL/6-
Tyr^c-2J-derived C2J ES cells. Correctly targeted ES cells were transiently transfected with a Flippase-puromycin plasmid to remove the PGK-neo-pA cassette. Neo-excised ES cells were injected into recipient blastocysts and chimeric males were bred with C57BL/6-Tyr^c-2J/J female mice. The donating investigator reports that the c-Kit^lox66-71 mice were crossed to C57BL/6; the albino allele (Tyr^c-2J)has been bred out.

Expression Data

Expressed Gene	GFP, Green Fluorescent Protein,
Site of Expression	EGFP is expressed in c-Kit-expressing cells.

Control Suggestions

Wild-type from the colony
000664 C57BL/6J

Additional Information

Considerations for Choosing Controls

Selected References

Kimura Y; Ding B; Imai N; Nolan DJ; Butler JM; Rafii S. 2011. c-Kit-Mediated Functional Positioning of Stem Cells to Their Niches Is Essential for Maintenance and Regeneration of Adult Hematopoiesis. PLoS One 6(10):e26918PubMed: 22046410 MGI: J:178071

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Genetics

Kit^tm1.1Sraf

Allele Symbol: Kit^tm1.1Sraf

Allele Name	targeted mutation 1.1, Shahin Rafii
Allele Type	Targeted (Reporter, Null/Knockout)
Allele Synonym(s)	c-Kit lox66-71
Gene Symbol and Name	Kit, KIT proto-oncogene receptor tyrosine kinase
Gene Synonym(s)
Expressed Gene	GFP, Green Fluorescent Protein,
Site of Expression	EGFP is expressed in c-Kit-expressing cells.
Strain of Origin	B6(Cg)-Tyr^c-2J/J
Chromosome	5
Molecular Note	A construct containing a lox66 site, inverted GFP gene, lox71, and FRT-flanked neo cassette was inserted between exon 8 and exon 9. Flp-mediated recombination removed the neo cassette.

Disease/Phenotype

Disease Terms

Research Areas By Phenotype

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

Research Tools
- Fluorescent Proteins

Mammalian Phenotype Terms by Genotype

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

Genotype: Kit^tm1.1Sraf/Kit⁺
involves: C57BL/6

mammalian phenotype

no phenotypic analysis

References

Kimura Y; Ding B; Imai N; Nolan DJ; Butler JM; Rafii S. 2011. c-Kit-Mediated Functional Positioning of Stem Cells to Their Niches Is Essential for Maintenance and Regeneration of Adult Hematopoiesis. PLoS One 6(10):e26918PubMed: 22046410 MGI: J:178071

Additional - Kit^tm1.1Sraf related

Technical Support

CONTACT TECHNICAL SUPPORT

Genotyping Protocols
- Pyrosequencing:Tyr
- Separated PCR:Kit-Alternate 4
- Sanger sequencing:Tyr-SEQ
- Genotyping resources and troubleshooting
Breeding Considerations

While maintaining a live colony, these mice are bred as heterozygotes. The donating investigator has not attempted to breed homozygous mice. (July 2016)
- Additional Breeding and Husbandry Support
Citation
When using the c-Kit^lox66-71 (c-Kit lox66-71) mouse strain in a publication, please cite the originating article(s) and include MMRRC stock #42035 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

The Jackson Laboratory's Genotype Promise

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. We do not guarantee breeding performance and therefore suggest that investigators order more than one breeding pair to avoid delays in their research.

Terms Of Use

General Terms and Conditions

See MMRRC for Additional Conditions of Distribution

Questions about Terms of Use

Licensing Information

Phone: 207-288-6470

Email: TechTran@jax.org

Also Known As:c-Kitlox66-71 (c-Kit lox66-71)

Donating Investigator

Genetic overview

Research Applications

Detailed Description

Development

Expression Data

Control Suggestions

Additional Information

Selected References

Kittm1.1Sraf

Allele Symbol: Kittm1.1Sraf

Disease Terms

Research Areas By Phenotype

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

Mammalian Phenotype Terms by Genotype

Genotype: Kittm1.1Sraf/Kit+involves: C57BL/6

mammalian phenotype

References

Additional - Kittm1.1Sraf related

Genotyping Protocols

Breeding Considerations

Citation

Animal Health Reports

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

The Jackson Laboratory's Genotype Promise

Terms Of Use

Licensing Information

Also Known As:c-Kit^lox66-71 (c-Kit lox66-71)

Kit^tm1.1Sraf

Allele Symbol: Kit^tm1.1Sraf

Genotype: Kit^tm1.1Sraf/Kit⁺
involves: C57BL/6

Additional - Kit^tm1.1Sraf related