A single-molecule method for the quantitation of microRNA gene expression
| Autor: | Mark Nadel, Joanne Garver, Jenny E. Rooke, Michael Gallo, Maria Hackett, Stephen McLaughlin, Lori Anne Neely, Steve Gullans, Sonal Patel, John Harris |
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| Rok vydání: | 2005 |
| Předmět: |
Oligonucleotide
Cell growth Cellular differentiation Gene Expression Profiling MicroRNA Gene Oligonucleotides Cell Biology Biology Oligonucleotides Antisense Non-coding RNA Biochemistry Molecular biology Sensitivity and Specificity Cell biology MicroRNAs microRNA Gene expression Humans Locked nucleic acid DNA Probes Molecular Biology In Situ Hybridization Biotechnology |
| Zdroj: | Nature methods. 3(1) |
| ISSN: | 1548-7091 |
| Popis: | MicroRNAs (miRNA) are short endogenous noncoding RNA molecules that regulate fundamental cellular processes such as cell differentiation, cell proliferation and apoptosis through modulation of gene expression. Critical to understanding the role of miRNAs in this regulation is a method to rapidly and accurately quantitate miRNA gene expression. Existing methods lack sensitivity, specificity and typically require upfront enrichment, ligation and/or amplification steps. The Direct miRNA assay hybridizes two spectrally distinguishable fluorescent locked nucleic acid (LNA)-DNA oligonucleotide probes to the miRNA of interest, and then tagged molecules are directly counted on a single-molecule detection instrument. In this study, we show the assay is sensitive to femtomolar concentrations of miRNA (500 fM), has a three-log linear dynamic range and is capable of distinguishing among miRNA family members. Using this technology, we quantified expression of 45 human miRNAs within 16 different tissues, yielding a quantitative differential expression profile that correlates and expands upon published results. |
| Databáze: | OpenAIRE |
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