Protocol abstract
Nature Protocols 3, - 563 - 578 (2008)
Published online: 13 March 2008 | doi:10.1038/nprot.2008.14
Subject Categories: Computational and theoretical biology | Genetic analysis | Genomics and proteomics | Nanotechnology | Nucleic acid based molecular biology
MicroRNA expression profiling using microarrays
Chang-Gong Liu1,4, George Adrian Calin1,3,4, Stefano Volinia1,2 & Carlo M Croce1
Abstract
Microarray technology is a powerful high-throughput tool capable of monitoring the expression of thousands of small noncoding RNAs at once within tens of samples processed in parallel in a single experiment. To conduct a genome-wide analysis of miRNA expression of normal and disease samples, such as cancer, and to distinguish expression signatures associated with diagnosis, prognosis and therapeutic interventions, we have developed a unique miRNA microarray assay on a CodeLink platform. The miRNA array consists of 4,104 probes printed in duplicate. This array can simultaneously profile more than 1,500 mature miRNAs and their corresponding precursors from 474 human and 373 mouse miRNA genes. The full protocol details of the miRNA microarray assay developed by our group are described here, including miRNA oligo probe design, array fabrication and miRNA target preparation (by reverse transcription of total RNA), target-probe hybridization on array, signal detection and data analysis. The assay is simple, can be easily standardized and allows the reproducible profiling of up to 24 total RNA samples within 24 h.
- Department of Molecular Virology, Immunology and Medical Genetics and Comprehensive Cancer Center, The Ohio State University, Columbus, Ohio 43210, USA.
- Morphology and Embryology Department, University of Ferrara, Ferrara 44100, Italy.
- Present address: Department of Experimental Therapeutics & Department of Cancer Genetics, University of Texas, MD Anderson Cancer Center, Houston, Texas 77030, USA.
- These authors contributed equally to this work.
Correspondence to: Carlo M Croce1 e-mail: Carlo.Croce@osumc.edu
nature-products
A-Z product listing
- 0.1 M dithiothreitol(Invitrogen)
- 0.22-
m Millipore sterile vacuum filter unit(Fisher) - 0.5 pmol l-1 of (5'-biotin-(dA)12-(dT-biotin)-(dA)12-(N)8-3') custom random octomer oligonucleotide primer for reverse transcription (RT) reaction(Sigma Genosys)
- 0.8-m Nalgene disposable filter unit(Fisher)
- 1 M Tris-HCl(Sigma)
- 1
PBS(Invitrogen)
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