C57BL/6-Egln1^tm1.1Fsl/J

Stock number: 023886
Research areas: Developmental Biology Research, Hematological Research, Research Tools

Description

These P294R knockin mice develop erythrocytosis and are useful in studies of erythropoiesis, the Prolyl Hydroxylase Domain protein/Hypoxia Inducible Factor pathway and physiological adaptation to hypoxic conditions.

Detailed description

The Egln1 gene encodes an oxygen sensing enzyme that regulates the activity of hypoxia inducible transcription factors (or HIF proteins) in normoxic and hypoxic conditions, enabling aerobic organisms to adapt to hypoxia. A nucleotide substitution mutation in the human gene, P317R, causes reduced enzymatic activity and has been linked to erythrocytosis. The mouse P294R mutation corresponds to the human P317R. Mice that are heterozygous for the P294R nucleotide substitution mutation are viable and fertile, but develop erythrocytosis. The mutation was confirmed by PCR analysis. Heterozygous mice develop erythrocytosis with increased spleen mass, increased red cell mass (increased hematocrit, hemoglobin, and red blood cell counts) but normal white blood cell and platelets numbers, and inappropriately normal serum levels of erythropoietin. Homozygotes are not viable.

Development

The C57BL/6 bacterial artificial chromosome (BAC) clone RP23-356I16 containing was modified by a minitargeting vector to insert the P249R nucleotide substitution. DNA from the modified BAC clone was used as the source for a targeting vector that also contained a loxP site flanked NEO cassette. This targeting vector was used to insert the P294R knockin mutation into exon 2. The construct was electroporated into C57BL/6 derived embryonic stem (ES) cells (Caliper Life Sciences). Correctly targeted ES cells were injected into C57BL/6 blastocysts. The resulting male chimeric animals were bred to C57BL/6 mice. The mice were then bred to C57BL/6-Gt(ROSA)26Sor^{tm16(Cre)Arte} mice to remove the selection cassette. Additional crossing with C57BL/6 mice was completed to breed out the Gt(ROSA)26Sor^{tm16(Cre)Arte} allele.

Upon arrival at The Jackson Laboratory, the mice were crossed to C57BL/6J (Stock No. 000664) at least once to establish the colony.

Allele

Symbol: Egln1^tm1.1Fsl
Name: targeted mutation 1.1, Frank Lee
MGI accession ID: MGI:5521374
Generation method: Targeted
Synonyms: Phd2^P294R
Marker symbol: Egln1
Marker name: egl-9 family hypoxia-inducible factor 1
Marker synonyms: C1orf12; ECYT3; HALAH; Hif-p4h-2; HIFPH2; HIF-PH2; HPH2; HPH-2; ORF13; PHD2; PHD-2; SM20; SM-20; ZMYND6
Chromosome: 8
Strain of origin: C57BL/6
Molecular note: The C57BL/6 bacterial artificial chromosome (BAC) clone RP23-356I16 containing genomic DNA from exons 2-4 of the Egln1 gene was modified to insert a P249R nucleotide substitution in exon 2 using a minitargeting vector. The final targeting vector contained a 6.9 kb 5' arm containing exon 2 with the P249R nucleotide substitution, a loxP-flanked neomycin selection cassette and a 4.1 kb 3' arm. The presence of the P249R mutation was confirmed by sequencing. The neomycin resistance cassette was removed by cre-mediated recombination using mice carrying the Gt(ROSA)26Sor^{tm16(cre)Arte} allele.