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Identification and detection of RNA-RNA interactions using the yeast RNA hybrid system

Nature Protocols volume 1, pages 689–694 (2006)Cite this article

Abstract

To test RNA-RNA interactions in cells, we developed a yeast RNA hybrid system derived from the yeast three-hybrid system. In this setup, the activation of a reporter gene (HIS3 or lacZ) is dependent on the interaction of two RNAs. One ('RNA X') is fused to MS2 RNA, forming the bait, which binds to a fusion protein composed of the MS2 coat and the LexA proteins. The second ('RNA Y') is fused to an RNA-based transcriptional activator (m26-11), forming the prey. If prey (RNA Y) binds to bait (RNA X), the m26-11 transcriptional activator is tethered to the promoter of the reporter genes. This protocol describes how to use this RNA hybrid system. In addition to testing RNA-RNA interactions, it can also be used to screen RNA libraries to identify new interaction partners or for mutational analysis of two known interaction partners.

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Figure 1: General description of the RNA hybrid system.
Figure 2: Library screening using the RNA hybrid system.

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Acknowledgements

Work in our laboratory is funded by the Austrian Science Fund (projects Z-72 and F1703), by the European community Research Programme Framework 6 (BACRNA FP6-018618) and by the Austrian BMBWK GenAU program.

Author information

Authors and Affiliations

  1. Institute of Biochemistry, University of Hamburg, Martin Luther King Platz 6, Hamburg, 20146, Germany

    Nicolas Piganeau

  2. Max F. Perutz Laboratories, University of Vienna, Dr. Bohrgasse 9/5, Vienna, A-1030, Austria

    Renée Schroeder

Authors
  1. Nicolas Piganeau
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  2. Renée Schroeder
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Contributions

N.P. developed the method and wrote most of this experimental procedure; R.S. is the supervisor and wrote the introduction and the anticipated results sections of the manuscript.

Corresponding author

Correspondence to Renée Schroeder.

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Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Figure 1

pIIIa m26 and pAN MS2-2. (PDF 25 kb)

Supplementary Data 1

pAN MS2-2. (TXT 9 kb)

Supplementary Data 2

pIIIa m26. (TXT 11 kb)

Supplementary Data 3

pAN X1 MS2. (TXT 9 kb)

Supplementary Data 4

pIIIa m26 Y1. (TXT 11 kb)

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Piganeau, N., Schroeder, R. Identification and detection of RNA-RNA interactions using the yeast RNA hybrid system. Nat Protoc 1, 689–694 (2006). https://doi.org/10.1038/nprot.2006.111

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