Subcellular Localization of MicroRNAs by MicroRNA In Situ Hybridization (miR-ISH)

Methods Mol Biol. 2019:2054:159-169. doi: 10.1007/978-1-4939-9769-5_11.

Abstract

MicroRNAs (miRNAs) are 22-nucleotide RNA sequences that regulate up to 60% of the mammalian transcriptome. Although canonical miRNA-induced silencing complex-mediated messenger RNA degradation occurs in the cytoplasm, miRNAs have been described in other subcellular compartments with potentially novel functions. Currently, there are limited methodologies for visualizing RNA locations within cells to elucidate mechanisms and pathways of miRNA biogenesis, transport, and function. Here, we describe a simple and rapid miRNA in situ hybridization method that can be combined with standard immunofluorescence procedures for subcellular localization of mature and precursor miRNAs.

Keywords: Hybridization; MicroRNA; Microscopy; RNA; Subcellular localization.

MeSH terms

  • Fluorescent Antibody Technique, Indirect / instrumentation
  • Fluorescent Dyes / chemistry
  • HEK293 Cells
  • Humans
  • In Situ Hybridization / instrumentation
  • In Situ Hybridization / methods*
  • Indoles / chemistry
  • MicroRNAs / metabolism*
  • Microscopy, Confocal / methods
  • Molecular Imaging / instrumentation
  • Molecular Imaging / methods*
  • Subcellular Fractions / metabolism

Substances

  • Fluorescent Dyes
  • Indoles
  • MicroRNAs
  • DAPI