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Live-cell imaging of Pol II promoter activity to monitor gene expression with RNA IMAGEtag reporters
Nucleic Acids Research
  • IIchung Shin, Iowa State University
  • Judhajeet Ray, Iowa State University
  • Vinayak Gupta, Iowa State University
  • Muslum IIgu, Iowa State University
  • Jonathan Beasley, Iowa State University
  • Lee Bendickson, Iowa State University
  • Samir Mehanovic, Iowa State University
  • George A. Kraus, Iowa State University
  • Marit Nilsen-Hamilton, Iowa State University
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We describe a ribonucleic acid (RNA) reporter system for live-cell imaging of gene expression to detect changes in polymerase II activity on individual promoters in individual cells. The reporters use strings of RNA aptamers that constitute IMAGEtags (Intracellular MultiAptamer GEnetic tags) that can be expressed from a promoter of choice. For imaging, the cells are incubated with their ligands that are separately conjugated with one of the FRET pair, Cy3 and Cy5. The IMAGEtags were expressed in yeast from the GAL1, ADH1 or ACT1 promoters. Transcription from all three promoters was imaged in live cells and transcriptional increases from the GAL1 promoter were observed with time after adding galactose. Expression of the IMAGEtags did not affect cell proliferation or endogenous gene expression. Advantages of this method are that no foreign proteins are produced in the cells that could be toxic or otherwise in- fluence the cellular response as they accumulate, the IMAGEtags are short lived and oxygen is not required to generate their signals. The IMAGEtag RNA reporter system provides a means of tracking changes in transcriptional activity in live cells and in real time.

This article is from Nucleic Acids Research; 2014, 42(11);e90. doi: 10.1093/nar/gku297. Posted with permission.

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IIchung Shin, Judhajeet Ray, Vinayak Gupta, Muslum IIgu, et al.. "Live-cell imaging of Pol II promoter activity to monitor gene expression with RNA IMAGEtag reporters" Nucleic Acids Research Vol. 42 Iss. 11 (2014) p. e90
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