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Functional Analysis of Noncoding RNAs in Trypanosomes: RNA Walk, a Novel Approach to Study RNA–RNA Interactions Between Small RNA and Its Target

  • Chaim Wachtel
  • Shulamit MichaeliEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 718)

Abstract

The recent discovery of thousands of small noncoding RNAs (ncRNAs), in many different organisms, has led to the need for methods to study their function. One way to help understand their function is to determine what other RNAs interact with the ncRNAs. We have developed a novel method to investigate the RNA–RNA interactions between a small RNA and its target that we termed “RNA walk.” The method is based on UV-induced AMT cross-linking in vivo followed by affinity selection of the hybrid molecules and mapping the intermolecular adducts by RT-PCR. Domains carrying the cross-linked adducts are less efficiently amplified than domains that are not cross-linked. Real-time PCR is used to quantify the results. Further mapping of the interactions is performed by primer extension to determine the exact cross-linked adduct.

Key words

RNA walk RNA–RNA interactions Cross-linking Affinity selection Trypanosomes 

Notes

Acknowledgement

This research was supported by a grant from the Israel–US Binational Science Foundation (BSF), and by an International Research Scholar’s Grant from the Howard Hughes Foundation to S.M. S.M. holds the David and Inez Myers Chair in RNA silencing of diseases.

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Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  1. 1.The Mina and Everard Goodman Faculty of Life Sciences and the Advanced Materials and Nanotechnology InstituteBar-Ilan UniversityRamat-GanIsrael

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