Direct and sensitive miRNA profiling from low-input total RNA.

RNA (New York, N.Y.) ·Vol. 13 ·No. 1 ·2007-01-00 ·Pages 151-9

Wang H, Ach RA, Curry B

Abstract

We have developed a sensitive, accurate, and multiplexed microRNA (miRNA) profiling assay that is based on a highly efficient labeling method and novel microarray probe design. The probes provide both sequence and size discrimination, yielding in most cases highly specific detection of closely related mature miRNAs. Using a simple, single-vial experimental protocol, 120 ng of total RNA is directly labeled using Cy3 or Cy5, without fractionation or amplification, to produce precise and accurate measurements that span a linear dynamic range from 0.2 amol to 2 fmol of input miRNA. The results can provide quantitative estimates of the miRNA content for the tissues studied. The assay is also suitable for use with formalin-fixed paraffin-embedded clinical samples. Our method allows rapid design and validation of probes for simultaneous quantitative measurements of all human miRNA sequences in the public databases and to new miRNA sequences as they are reported.

MeSH Terms

Animals Carbocyanines/chemistry Gene Expression Profiling/methods Humans MicroRNAs/analysis Oligonucleotide Array Sequence Analysis/methods RNA Probes/chemistry Sensitivity and Specificity Tissue Distribution

Chemicals

Carbocyanines MicroRNAs RNA Probes cyanine dye 3 cyanine dye 5

Authors & Affiliations

3 authors, click to expand affiliations / ORCID

Wang Hui

Agilent Technologies, Inc., Agilent Laboratories, Santa Clara, California 95051, USA. hui_wang@agilent.com

Ach Robert A

Curry Bo

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Article Info

Journal

RNA (New York, N.Y.)

Abbr.

RNA

ISSN

1355-8382

DOI

10.1261/rna.234507

hui_wang@agilent.com

Published

2007-01-00

Epub

2006-00-14

Pages

151-9

Language

English

Region

United States

NLM ID

9509184

PMCID

PMC1705746

Subset

External Links

PubMed source DOI: 10.1261/rna.234507 PMC Full Text View journal details

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