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http://dx.doi.org/10.5483/BMBRep.2010.43.2.110

On-off controllable RNA hybrid expression vector for yeast three-hybrid system  

Bak, Geunu (Department of Chemistry, KAIST)
Hwang, Se-Won (Korea Academy of Science of KAIST)
Ko, Ye-Rim (Department of Chemistry, KAIST)
Lee, Jung-Min (Department of Chemistry, KAIST)
Kim, Young-Mi (Department of Chemistry, KAIST)
Kim, Kyung-Hwan (Department of Chemistry, KAIST)
Hong, Soon-Kang (Department of Fire Service Administration, Chodang University)
Lee, Young-Hoon (Department of Chemistry, KAIST)
Publication Information
BMB Reports / v.43, no.2, 2010 , pp. 110-114 More about this Journal
Abstract
The yeast three-hybrid system (Y3H), a powerful method for identifying RNA-binding proteins, still suffers from many false positives, due mostly to RNA-independent interactions. In this study, we attempted to efficiently identify false positives by introducing a tetracycline operator (tetO) motif into the RPR1 promoter of an RNA hybrid expression vector. We successfully developed a tight tetracycline-regulatable RPR1 promoter variant containing a single tetO motif between the transcription start site and the A-box sequence of the RPR1 promoter. Expression from this tetracycline-regulatable RPR1 promoter in the presence of tetracycline-response transcription activator (tTA) was positively controlled by doxycycline (Dox), a derivative of tetracycline. This on-off control runs opposite to the general knowledge that Dox negatively regulates tTA. This positively controlled RPR1 promoter system can therefore efficiently eliminate RNA-independent false positives commonly observed in the Y3H system by directly monitoring RNA hybrid expression.
Keywords
RNA hybrid expression vector; RPR1 promoter; Tetracycline regulation; Yeast three-hybrid system;
Citations & Related Records
Times Cited By KSCI : 1  (Citation Analysis)
Times Cited By Web Of Science : 1  (Related Records In Web of Science)
Times Cited By SCOPUS : 3
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