Elk-1 knock-out mice engineered by Flp recombinase-mediated cassette exchange.
Elk-1 is a member of the TCF subfamily of Ets proteins. TCFs interact with SRF at serum response elements (SREs) of immediate early genes (IEGs), such as c-fos and Egr-1, thereby mediating IEG induction upon extracellular stimulation. We previously generated an Elk-1 null allele (Elk1-137) in murine embryonic stem (ES) cells by homologous recombination. In Elk1-137, the Elk-1 gene was replaced by a Hygromycin B phosphotransferase - Thymidine Kinase (HygTk) fusion gene, flanked by two nonidentical Flp recombinase recognition (FRT) sites (Cesari et al., [2004] Mol Cell Biol, in press) to allow for the subsequent generation of alternative alleles of interest by recombinase-mediated cassette exchange (RMCE). Elk1-deficient mice derived from Elk-1((137/0)) ES cells are viable and do not reveal strong phenotypical abnormalities, apart from male sterility. However, the Elk-1 locus contains the Tk cassette, which has previously been related to this defect. Therefore, in our first experiment involving the technique of Flp RMCE we chose to remove the HygTk cassette in Elk-1((137/0)) ES cells and to generate Elk-1((RMCE16/0)) and Elk-1((RMCE16/RMCE16)) mice. In so doing, we provide evidence that the sterility of Elk1((137/0)) mice was not due to the absence of Elk-1 but rather the presence of HygTk. This is the first report of mice derived from ES cells which were subjected to Flp RMCE and thus proves that RMCE is a powerful tool for the genetic engineering of previously tagged loci in the mouse genome.
Cesari
Francesca
F
Rennekampff
Verena
V
Vintersten
Kristina
K
Vuong
Lam Giang
LG
Seibler
Jost
J
Bode
Jürgen
J
Wiebel
Franziska F
FF
Nordheim
Alfred
A
D000818:Animals; D004254:DNA Nucleotidyltransferases / Q000235:genetics* / Q000378:metabolism; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D017353:Gene Deletion; D017781:Genes, Immediate-Early / Q000235:genetics; D005818:Genetic Engineering / Q000379:methods*; D051379:Mice; D018345:Mice, Knockout; D016254:Mutagenesis, Insertional / Q000379:methods; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics*; D016133:Polymerase Chain Reaction; D011518:Proto-Oncogene Proteins / Q000235:genetics* / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D026362:Serum Response Factor / Q000378:metabolism; D013937:Thymidine Kinase / Q000235:genetics*; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism; D050807:ets-Domain Protein Elk-1
D017931:DNA Primers; D004268:DNA-Binding Proteins; C494113:Elk1 protein, mouse; D011518:Proto-Oncogene Proteins; D026362:Serum Response Factor; D014157:Transcription Factors; D050807:ets-Domain Protein Elk-1; D017853:Phosphotransferases (Alcohol Group Acceptor); C063242:hygromycin-B kinase; D013937:Thymidine Kinase; D004254:DNA Nucleotidyltransferases; C045119:FLP recombinase