Reporter-linked monitoring of transgene expression in living cells using the ecdysone-inducible promoter system.
Inducible promoter systems such as the ecdysone-inducible system or the tetracycline-regulated expression systems have proven to be powerful tools in studying gene function. In practice, such systems have met with the difficulty that either the vector expressing the transactivator gene or the vector carrying the response element are frequently silenced by flanking genomic sequences after stable integration. In order to identify those cells in a heterogeneous population in which a transgene is expressed from an ecdysone-inducible promoter, we have created the vector p2ER-EGFP/mcs that contains two ecdysone-inducible expression cassettes in tandem. Using two reporter genes, lacZ and green fluorescent protein (EGFP), we demonstrate that the expression of both genes can be co-induced from a very low baseline in CHO cells expressing the modified ecdysone receptor and the retinoid X receptor. The expression of EGFP and lacZ from vector p2ER-EGFP/lacZ follows the same Muristerone A concentration-dependence as that of EGFP from vector pER-EGFP, indicating that the juxtaposition of the two inducible promoters in vector p2ER-EGFP/mcs does not cause cross interference between them. We suggest that this modification of the ecdysone-inducible promoter system will allow for the visual control of the induced expression of other genes by Muristerone A.
Lüers
G H
GH
University of Bonn, Institute for Anatomy, Germany.
Jess
N
N
Franz
T
T
D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D004440:Ecdysone / Q000235:genetics*; D004441:Ecdysterone / Q000031:analogs & derivatives* / Q000235:genetics; D005786:Gene Expression Regulation / Q000502:physiology; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins; D007202:Indicators and Reagents / Q000378:metabolism; D008164:Luminescent Proteins / Q000235:genetics; D008967:Molecular Biology / Q000379:methods*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012333:RNA, Messenger / Q000032:analysis; D014162:Transfection; D019076:Transgenes / Q000502:physiology*
D007202:Indicators and Reagents; D008164:Luminescent Proteins; D012333:RNA, Messenger; D049452:Green Fluorescent Proteins; D004440:Ecdysone; C055348:muristerone A; D004441:Ecdysterone