Generation of inducible Cre systems for conditional gene inactivation in mice

摘要: Inducible gene targeting based on the Cre/loxP system is a novel technique allowing conditional modification in mice. Precondition of this method regulation Cre activity by inducible genetic switches. This study aimed at improving systems for both transcriptional and posttranslational level, which should be tested first vitro subsequently newly generated transgenic In order to detect activity, special double indicator consisting DsRed eGFP was constructed targeted into ROSA26 locus HM-1 ES cells. constitutively express flanked loxP sites. Cre-mediated recombination excises turns gene, as evidenced change fluorescence from red green. Though worked properly transiently transfected fibroblasts, strategy used integration presumably disrupted putative stem loop motif 5�UTR resulting silencing protein level. However, cell line with integrated suited targetings constructs HPRT locus, since inducibilities are detectable Southern Blot analysis. The Cre*PR one I have established. contrast classical CrePR1 system, background almost abolished introducing point mutation 3�end part, thus preventing aberrant splicing encoding mRNA. Also, PR-LBD improved, compared CrePR1, obtain higher affinity towards inducer RU486. driven either EF1£\ promoter or CAGGS promoter, then locus. cells expected. These were generate these mice, high levels obtained achieve excision loxP-flanked segments. addition, B specific mouse strain mCD19 another approach inducibly regulate Cre, tet-on established rtTA-M2 transactivator tetresponsive cassette, line. revealed tight expression doxycycline dependent manner.