Document Detail

Quick Fluorescent In Situ Hybridization Protocol for Xist RNA Combined with Immunofluorescence of Histone Modification in X-chromosome Inactivation.
MedLine Citation:
PMID:  25489864     Owner:  NLM     Status:  In-Data-Review    
Combining RNA fluorescent in situ hybridization (FISH) with immunofluorescence (immuno-FISH) creates a technique that can be employed at the single cell level to detect the spatial dynamics of RNA localization with simultaneous insight into the localization of proteins, epigenetic modifications and other details which can be highlighted by immunofluorescence. X-chromosome inactivation is a paradigm for long non-coding RNA (lncRNA)-mediated gene silencing. X-inactive specific transcript (Xist) lncRNA accumulation (called an Xist cloud) on one of the two X-chromosomes in mammalian females is a critical step to initiate X-chromosome inactivation. Xist RNA directly or indirectly interacts with various chromatin-modifying enzymes and introduces distinct epigenetic landscapes to the inactive X-chromosome (Xi). One known epigenetic hallmark of the Xi is the Histone H3 trimethyl-lysine 27 (H3K27me3) modification. Here, we describe a simple and quick immuno-FISH protocol for detecting Xist RNA using RNA FISH with multiple oligonucleotide probes coupled with immunofluorescence of H3K27me3 to examine the localization of Xist RNA and associated epigenetic modifications. Using oligonucleotide probes results in a shorter incubation time and more sensitive detection of Xist RNA compared to in vitro transcribed RNA probes (riboprobes). This protocol provides a powerful tool for understanding the dynamics of lncRNAs and its associated epigenetic modification, chromatin structure, nuclear organization and transcriptional regulation.
Minghui Yue; John Lalith Charles Richard; Norishige Yamada; Akiyo Ogawa; Yuya Ogawa
Related Documents :
8401494 - Digital dna typing at a second hypervariable locus by minisatellite variant repeat mapp...
7907444 - Female twins with severe christmas disease (hemophilia b).
9770504 - Orientation-dependent and sequence-specific expansions of ctg/cag trinucleotide repeats...
15317214 - Molecular genetic data at two tetranucleotide repeat loci (d12s66 and d12s67) in two in...
12551824 - Cytogenetic and molecular delineation of a region of chromosome 3q commonly gained in m...
3477874 - Identification of nucleotide-excision-repair genes on human chromosomes 2 and 13 by fun...
Publication Detail:
Type:  Journal Article     Date:  2014-11-26
Journal Detail:
Title:  Journal of visualized experiments : JoVE     Volume:  -     ISSN:  1940-087X     ISO Abbreviation:  J Vis Exp     Publication Date:  2014  
Date Detail:
Created Date:  2014-12-10     Completed Date:  -     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  101313252     Medline TA:  J Vis Exp     Country:  United States    
Other Details:
Languages:  eng     Pagination:  -     Citation Subset:  IM    
Export Citation:
APA/MLA Format     Download EndNote     Download BibTex
MeSH Terms

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine

Previous Document:  How Accurately Can We Predict the Melting Points of Drug-like Compounds?
Next Document:  [GINA 2014: Yin and Yang].