title: Whole mount RNA fluorescent in situ hybridization of Drosophila embryos.
abstract: Assessing the expression pattern of a gene, as well as the subcellular localization properties of its transcribed RNA, are key features for understanding its biological function during development. RNA in situ hybridization (RNA-ISH) is a powerful method used for visualizing RNA distribution properties, be it at the organismal, cellular or subcellular levels. RNA-ISH is based on the hybridization of a labeled nucleic acid probe (e.g. antisense RNA, oligonucleotides) complementary to the sequence of an mRNA or a non-coding RNA target of interest. As the procedure requires primary sequence information alone to generate sequence-specific probes, it can be universally applied to a broad range of organisms and tissue specimens. Indeed, a number of large-scale ISH studies have been implemented to document gene expression and RNA localization dynamics in various model organisms, which has led to the establishment of important community resources. While a variety of probe labeling and detection strategies have been developed over the years, the combined usage of fluorescently-labeled detection reagents and enzymatic signal amplification steps offer significant enhancements in the sensitivity and resolution of the procedure. Here, we describe an optimized fluorescent in situ hybridization method (FISH) employing tyramide signal amplification (TSA) to visualize RNA expression and localization dynamics in staged Drosophila embryos. The procedure is carried out in 96-well PCR plate format, which greatly facilitates the simultaneous processing of large numbers of samples.
authors: Institut de Recherches Cliniques de Montréal.
authors: Félix
authors: F
authors: Legendre
authors: Neal
authors: N
authors: Cody
authors: Carole
authors: C
authors: Iampietro
authors: Julie
authors: J
authors: Bergalet
authors: Fabio Alexis
authors: FA
authors: Lefebvre
authors: Gaël
authors: G
authors: Moquin-Beaudry
authors: Olivia
authors: O
authors: Zhang
authors: Xiaofeng
authors: X
authors: Wang
authors: Eric
authors: E
authors: Lécuyer
mesh_terms: D000818:Animals; D004330:Drosophila / Q000737:chemistry / Q000196:embryology / Q000235:genetics*; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*; D012313:RNA / Q000032:analysis* / Q000096:biosynthesis / Q000235:genetics
chemical_list: D012313:RNA