title: Small RNA Detection by in Situ Hybridization Methods.
abstract: Small noncoding RNAs perform multiple regulatory functions in cells, and their exogenous mimics are widely used in research and experimental therapies to interfere with target gene expression. MicroRNAs (miRNAs) are the most thoroughly investigated representatives of the small RNA family, which includes short interfering RNAs (siRNAs), PIWI-associated RNA (piRNAs), and others. Numerous methods have been adopted for the detection and characterization of small RNAs, which is challenging due to their short length and low level of expression. These include molecular biology methods such as real-time RT-PCR, northern blotting, hybridization to microarrays, cloning and sequencing, as well as single cell miRNA detection by microscopy with in situ hybridization (ISH). In this review, we focus on the ISH method, including its fluorescent version (FISH), and we present recent methodological advances that facilitated its successful adaptation for small RNA detection. We discuss relevant technical aspects as well as the advantages and limitations of ISH. We also refer to numerous applications of small RNA ISH in basic research and molecular diagnostics.
authors.affiliation: Department of Molecular Biomedicine, Institute of Bioorganic Chemistry, Polish Academy of Sciences, Noskowskiego 12/14 Str., 61-704 Poznan, Poland. martyna.urbanek@ibch.poznan.pl.
authors.forename: Martyna O
authors.initials: MO
authors.lastname: Urbanek
authors.affiliation: Department of Molecular Biomedicine, Institute of Bioorganic Chemistry, Polish Academy of Sciences, Noskowskiego 12/14 Str., 61-704 Poznan, Poland. anna.u.nawrocka@gmail.com.
authors.forename: Anna U
authors.initials: AU
authors.lastname: Nawrocka
authors.affiliation: Department of Molecular Biomedicine, Institute of Bioorganic Chemistry, Polish Academy of Sciences, Noskowskiego 12/14 Str., 61-704 Poznan, Poland. wlodkrzy@ibch.poznan.pl.
authors.forename: Wlodzimierz J
authors.initials: WJ
authors.lastname: Krzyzosiak
mesh_terms: D000818:Animals; D006801:Humans; D017403:In Situ Hybridization / Q000379:methods*; D035683:MicroRNAs / Q000737:chemistry* / Q000235:genetics; D057089:Pathology, Molecular / Q000379:methods
chemical_list: D035683:MicroRNAs
keywords: LNA probe; PLA; Piwi-interacting RNA; TIRCA; enzyme-labeled fluorescence signal amplification; padlock probes; rolling circle amplification; short interfering RNA; tyramide signal amplification