C57BL/6-Gt(ROSA)26Sor^{tm8(Map2k1*,EGFP)Rsky}/J

Stock number: 012352
Research areas: Cancer Research, Cell Biology Research, Immunology, Inflammation and Autoimmunity Research, Research Tools

Description

The R26Stop^FLMEK1DD conditional allele is targeted to the Gt(ROSA)26Sor locus and has a loxP-flanked Neo-STOP cassette preventing transcription of the downstream bicistronic sequences (MEK1DD [a mutant form of rat MAPKK1 rendered constitutively active by two serine->aspartic acid substitutions (S218D/S222D) within the catalytic domain] and EGFP). These R26Stop^FLMEK1DD mice allow inducible constitutive activity of MAP Kinase signal transduction pathways that mediate various cellular activities, including gene expression, mitosis, differentiation, proliferation, cell survival, and cell cycle progression.

Detailed description

Mice homozygous for the R26Stop^FLMEK1DD conditional allele are viable and fertile, with a loxP-flanked Neo-STOP cassette preventing transcription of the downstream bicistronic sequences (MEK1DD [a mutant form of rat MAPKK1 rendered constitutively active by two serine->aspartic acid substitutions (S218D/S222D) within the catalytic domain] and EGFP). When bred to mice that express Cre recombinase, offspring will have the STOP cassette deleted and subsequent expression of the MEK1DD signal molecule and EGFP fluorescence in the cre-expressing cells. Expression of MEK1DD leads to constitutive activity of MAP Kinase signal transduction pathways that mediate various cellular activities, including gene expression, mitosis, differentiation, proliferation, cell survival, and cell cycle progression. Of note, breeding these mice to an FLP-expressing strain will result in removal of the frt-flanked IRES-EGFP cassette.

Development

A targeting vector designed with (from 5' to 3') a splice acceptor site, a loxP-flanked Neo-STOP cassette, a cDNA sequence encoding rat MEK1DD (a mutant form of MAPKK1 rendered constitutively active by two serine->aspartic acid substitutions (S218D/S222D) within the catalytic domain], an frt-flanked internal ribosomal entry site-enhanced green fluorescent protein cassette (IRES-EGFP), and bovine polyadenylation signal. This transgene construct was inserted between exons 1 and 2 of the Gt(ROSA)26Sor locus via electroporation into C57BL/6-derived embryonic stem (ES) cells. Correctly targeted ES cells were selected and chimeric animals were bred to C57BL/6 to generate the mutant colony. The donating investigator reports that these R26Stop^FLMEK1DD mice were maintained on a C57BL/6 genetic background (see SNP note below) as a homozygous colony for many generations prior to arrival at The Jackson Laboratory Repository. Upon arrival, mutant mice were bred to C57BL/6J inbred mice (Stock No. 000664) for at least one generation to establish the colony.

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. In some of the mice, one marker on chromosome 11 (~8.4 Mbp) and one marker on chromosome 17 (~34.3 Mbp) were not fixed to the C57BL/6 genetic background. The source of this is not known. In addition, at least 3 of 5 markers that determine C57BL/6J from C57BL/6N were found to be segregating in all the rederived mice. These data suggest the mice sent to The Jackson Laboratory Repository were on a mixed C57BL/6J ; C57BL/6N genetic background.

Allele

Symbol: Gt(ROSA)26Sor^{tm8(Map2k1*,EGFP)Rsky}
Name: targeted mutation 8, Klaus Rajewsky
MGI accession ID: MGI:4430550
Generation method: Targeted
Attributes: Conditional ready (e.g. floxed); Reporter; Constitutively active; Inserted expressed sequence
Marker symbol: Gt(ROSA)26Sor
Marker name: gene trap ROSA 26, Philippe Soriano
Chromosome: 6
Strain of origin: C57BL/6
Expressed genes: Map2k1,mitogen activated protein kinase kinase 1,rat; GFP,Green Fluorescent Protein,
Site of expression: Cre excision of the stop signal results in expression of EGFP and a constitutively active form of rat MAPKK1 (MEK1DD) in cre-expressing tissues.
Molecular note: A targeting vector designed with (from 5' to 3') a splice acceptor site, a loxP-flanked Neo-STOP cassette, a cDNA sequence encoding rat MEK1DD (a mutant form of MAPKK1 rendered constitutively active by two serine to aspartic acid substitutions (S218D/S222D) within the catalytic domain], an FRT-flanked internal ribosomal entry site-enhanced green fluorescent protein cassette (IRES-EGFP), and bovine polyadenylation signal. This transgene construct was inserted between exons 1 and 2 of the Gt(ROSA)26Sor locus.