STOCK Col1a1^{tm1.1(tetO-Gfi1,-Venus)Hlg}/J

Stock number: 032712
Product name: Gfi1-IRES-Venus (G3GV) knock-in
Research areas: Neurobiology Research, Research Tools

Description

The G3GV (G3- Gfi1-IRES-Venus eYFP) strain carries a doxycycline-inducible Col1a1 tetO-Gfi1-IRES-Venus knock-in allele. When bred with another transgenic mouse expressing reverse tetracycline-controlled transactivator protein (rtTA) or tetracycline-controlled transactivator protein (tTA), Cre mediated recombination in the resulting bitransgenic offspring can be regulated with the tetracycline analog, doxycycline. These mice may be suitable generating inducible, tissue-specific, targeted mutants useful in studies related to granulopoietic differentiation.

Detailed description

These G3GV (G3- Gfi1-IRES-Venus eYFP) mice express doxycycline-inducible mouse Gfi1-IRES-Venus. G3GV mice carry the doxycycline-inducible Gfi1-IRES-Venus under the control of the tetO, tetracycline-responsive regulatory element. The Gfi1 gene encodes a transcription factor involved in granulocyte specification (granulopoiesis) and restricts proliferation of hematopoietic stem cells. When these transgenic mice are bred with other transgenic mice expressing reverse tetracycline-controlled transactivator protein (rtTA) or tetracycline-controlled transactivator protein (tTA), tissue-specific Gfi1-IRES-Venus expression can be regulated with the tetracycline analog, doxycycline. Homozygotes are viable and fertile.

When crossed to mice carrying the Gt(ROSA)26Sor^{tm1(rtTA*M2)Jae} allele, doxycycline treatment of the resulting double mutant mice induces repression of hematopoietic stem cells while promoting granulopoietic differentiation, as well as Venus eYFP expression in granulocyte monocyte progenitor cells. The Donating Investigator notes that prolonged doxycycline administration results in gastrointestinal swelling and death. In the absence of the tetracycline analog doxycycline (dox), no phenotype is observed.

Development

A targeting vector containing sequence encoding the mouse Gfi1 open reading frame, followed by IRES-Venus (YFP variant) under the control of the tetO (tetracycline responsive element) was utilized in the construction of this mutant line. The targeting vector was electroporated into KH2 embryonic stem (ES) cells, which carry the Gt(ROSA)26Sor^{tm1(rtTA*M2)Jae} allele. Correctly targeted ES cells, carrying Gt(ROSA)26Sor^{tm1(rtTA*M2)Jae} and with Gfi1-IRES-Venus inserted into the 3' UTR of the Col1a1 locus, were transiently transfected with a FLP recombinase expression vector (pCAGs-FLPe-Puro), and then injected into blastocysts. The resulting chimeric animals were tested for germline transmission. The mice were bred to mutant lines carrying rtTA3 and Cre recombinase alleles. Upon arrival, sperm was cryopreserved. To establish our live colony, an aliquot of frozen sperm was used to fertilize C57BL/6J oocytes (Stock No. 000664), the rtTA3 and Cre recombinase alleles were bred out and sperm from mice carrying only the Gfi1-IRES-Venus allele was cryopreserved.



In 2019, a 48 SNP (single nucleotide polymorphism) panel analysis, with 43 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. One of the markers on chromosome 10 was segregating with an unknown source and one marker on chromosome 12 was segregating with 129. Also, one of the 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/6N;C57BL/6J genetic background.

Allele

Symbol: Col1a1^{tm1.1(tetO-Gfi1,-Venus)Hlg}
Name: targeted mutation 1.1, H Leighton Grimes
MGI accession ID: MGI:6281947
Generation method: Targeted
Attributes: Reporter; Inducible; Inserted expressed sequence
Synonyms: G3GV; Gfi1-IRES-Venus
Marker symbol: Col1a1
Marker name: collagen, type I, alpha 1
Marker synonyms: CAFYD; Col1a-1; Cola1; Cola-1; COLIA1; EDSARTH1; EDSC; Mov-13; OI1; OI2; OI3; OI4
Chromosome: 11
Strain of origin: (C57BL/6 x 129S4/SvJae)F1
Molecular note: The targeting vector contains sequence encoding the mouse Gfi1 open reading frame, followed by an IRES-Venus (YFP variant) under the control of the tetO (tetracycline responsive element). The Gfi1-IRES-Venus vector is inserted into the 3' UTR of the Col1a1 locus using a FLP recombinase expression vector (pCAGs-FLPe-Puro), which also repairs a defective hygromycin resistance gene.
General note: The allele was made in KH2 cells that carry Gt(ROSA)26Sor^{tm1(rtTA*M2)Jae} and Col1a1^{tm2(tetO-Pou5f1)Jae}.