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Detection of Small Noncoding RNAs by In Situ Hybridization Using Probes of 2′-O-Methyl RNA + LNA

  • Martin Jensen Søe
  • Martin Dufva
  • Kim HolmstrømEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1173)

Abstract

In situ hybridization is a powerful method to provide information about contextual distribution and cellular origin of nucleic acids, e.g., in formalin-fixed paraffin-embedded (FFPE) samples of tissue. Particularly the recently discovered classes of noncoding RNA (ncRNA) including endo-siRNAs and microRNAs require such a technique to enable their study and visualization in natural contexts, and in the last decade, many advances have been made, increasing our ability to specifically detect small ncRNAs. One of the key developments has been the demonstration of the superiority of using locked nucleic acid (LNA)-modified DNA probes for the detection of ncRNA in tissue. Here, we describe an alternative in situ hybridization protocol employing oligonucleotide probes consisting of combinations of LNA and 2´-O-methyl RNAs that under optimized hybridization buffer conditions can provide a highly sensitive assay performance with only 1 h hybridization time.

Keywords

Noncoding RNA microRNA LNA In situ hybridization 2′-O-methyl RNA 

Notes

Acknowledgments

The authors thank the Danish Ministry and Agency of Science, Technology, and Innovation for funding to MJS.

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Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  • Martin Jensen Søe
    • 1
  • Martin Dufva
    • 2
  • Kim Holmstrøm
    • 1
    Email author
  1. 1.Bioneer A/SHørsholmDenmark
  2. 2.DTU NanotechTechnical University of DenmarkLyngbyDenmark

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