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A sensitive alternative for microRNA in situ hybridizations using probes of 2′-o-methyl RNA + LNA

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  • Martin Jensen Søe
  • Trine Møller
  • Martin Dufva
  • Kim Holmstrøm
The use of short, high-affinity probes consisting of a combination of DNA and locked nucleic acid (LNA) has enabled the
specific detection of microRNAs (miRNAs) by in situ hybridization (ISH). However, detection of low–copy number miRNAs
is still not always possible. Here the authors show that probes consisting of 2′-O-methyl RNAs (2OMe) and LNA at every
third base (2:1 ratio), under optimized hybridization conditions, excluding yeast RNA from the hybridization buffer, can
provide superior performance in detection of miRNA targets in terms of sensitivity and signal-to-noise ratio compared
to DNA + LNA probes. Furthermore, they show that hybridizations can be performed in buffers of 4M urea instead of
50% formamide, thereby yielding an equally specific but nontoxic assay. The use of 2OMe + LNA–based probes and the
optimized ISH assay enable simple and fast detection of low–copy number miRNA targets, such as miR-130a in mouse brain.
OriginalsprogEngelsk
TidsskriftJournal of Histochemistry and Cytochemistry
Vol/bind59
Udgave nummer7
Sider (fra-til)661-672
Antal sider12
ISSN0022-1554
DOI
StatusUdgivet - 2011
Eksternt udgivetJa

ID: 110338926