Application of an RNA amplification method for reliable single-cell transcriptome analysis

Application of an RNA amplification method for reliable single-cell transcriptome analysis

Diverse cell types have unique transcriptional signatures that are best interrogated at single-cell resolution. Here we describe a novel RNA amplification approach that allows for high fidelity gene profiling of individual cells. This technique significantly diminishes the problem of 3' bias, e...

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Veröffentlicht in:BioTechniques. - 1993. - 59(2015), 3 vom: 08. Sept., Seite 137-48
1. Verfasser: Suslov, Oleg (VerfasserIn)
Weitere Verfasser: Silver, Daniel J, Siebzehnrubl, Florian A, Orjalo, Arturo, Ptitsyn, Andrey, Steindler, Dennis A
Format: Online-Aufsatz
Sprache:English
Veröffentlicht: 2015
Zugriff auf das übergeordnete Werk:BioTechniques
Schlagworte:Journal Article Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't RNA amplification SVZ single-cell analysis stem and progenitor cells AC133 Antigen Antigens, CD DNA-Binding Proteins mehr... Eye Proteins Glial Fibrillary Acidic Protein Glycoproteins Homeodomain Proteins Idb1 protein, mouse Inhibitor of Differentiation Protein 1 Membrane Proteins Nerve Tissue Proteins NeuN protein, mouse Nuclear Proteins Numb protein, mouse PAX6 Transcription Factor PAX6 protein, human Paired Box Transcription Factors Pax6 protein, mouse Peptides Repressor Proteins glial fibrillary astrocytic protein, mouse Green Fluorescent Proteins 147336-22-9 RNA 63231-63-0 EGFR protein, mouse EC 2.7.10.1 ErbB Receptors
Beschreibung
Zusammenfassung:Diverse cell types have unique transcriptional signatures that are best interrogated at single-cell resolution. Here we describe a novel RNA amplification approach that allows for high fidelity gene profiling of individual cells. This technique significantly diminishes the problem of 3' bias, enabling detection of all regions of transcripts, including the recognition of mRNA with short or completely absent poly(A) tails, identification of noncoding RNAs, and discovery of the full array of splice isoforms from any given gene product. We assess this technique using statistical and bioinformatics analyses of microarray data to establish the limitations of the method. To demonstrate applicability, we profiled individual cells isolated from the mouse subventricular zone (SVZ)-a well-characterized, discrete yet highly heterogeneous neural structure involved in persistent neurogenesis. Importantly, this method revealed multiple splice variants of key germinal zone gene products within individual cells, as well as an unexpected coexpression of several mRNAs considered markers of distinct and separate SVZ cell types. These findings were independently confirmed using RNA-fluorescence in situ hybridization (RNA-FISH), contributing to the utility of this new technology that offers genomic and transcriptomic analysis of small numbers of dynamic and clinically relevant cells
Beschreibung:Date Completed 02.06.2016
Date Revised 10.12.2019
published: Electronic-eCollection
Citation Status MEDLINE
ISSN:1940-9818
DOI:10.2144/000114331