Abstract
MicroRNAs (miRNAs) are important cellular modulators that regulate gene expression at the posttranscriptional level. Circulating miRNAs present in human plasma or serum have recently become an emerging field in biomedical research, mostly due to its potential applications in the diagnosis and prognosis of several diseases. Although miRNA profiling in biofluids holds great promise, there are several challenges to overcome. Here, we present an experimental procedure for profiling miRNA in plasma or serum with high sensitivity and specificity using qRT-PCR. This method is also suitable for studying miRNAs in other body fluids or clinical samples that also contain low amounts of RNA.
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References
Bartel DP (2009) MicroRNAs: target recognition and regulatory functions. Cell 136: 215–233
Zhang S, Chen L, Jung EJ et al (2010) Targeting microRNAs with small molecules: from dream to reality. Clin Pharmacol Ther 87:754–758
Jung M, Schaefer A, Steiner I et al (2010) Robust microRNA stability in degraded RNA preparations from human tissue and cell samples. Clin Chem 56:998–1006
Brase JC, Johannes M, Schlomm T et al (2011) Circulating miRNAs are correlated with tumor progression in prostate cancer. Int J Cancer 128:608–616
Yu DC, Li QG, Ding XW et al (2011) Circulating MicroRNAs: potential biomarkers for cancer. Int J Mol Sci 12:2055–2063
D’Alessandra Y, Devanna P, Limana F et al (2010) Circulating microRNAs are new and sensitive biomarkers of myocardial infarction. Eur Heart J 31:2765–2773
Dimmeler S, Zeiher AM (2010) Circulating microRNAs: novel biomarkers for cardiovascular diseases? Eur Heart J 31:2705–2707
Chen X, Hu Z, Wang W et al (2012) Identification of ten serum microRNAs from a genome-wide serum microRNA expression profile as novel noninvasive biomarkers for nonsmall cell lung cancer diagnosis. Int J Cancer 130:1620–1628
Gilad S, Meiri E, Yogev Y et al (2008) Serum microRNAs are promising novel biomarkers. PLoS One 3:e3148
Lawrie CH, Gal S, Dunlop HM et al (2008) Detection of elevated levels of tumour-associated microRNAs in serum of patients with diffuse large B-cell lymphoma. Br J Haematol 141: 672–675
Fichtlscherer S, De Rosa S, Fox H et al (2010) Circulating microRNAs in patients with coronary artery disease. Circ Res 107: 677–684
Komatsu S, Ichikawa D, Takeshita H et al (2011) Circulating microRNAs in plasma of patients with oesophageal squamous cell carcinoma. Br J Cancer 105:104–111
Wang GK, Zhu JQ, Zhang JT et al (2010) Circulating microRNA: a novel potential biomarker for early diagnosis of acute myocardial infarction in humans. Eur Heart J 31: 659–666
Blondal T, Jensby NS, Baker A et al (2013) Assessing sample and miRNA profile quality in serum and plasma or other biofluids. Methods 59:S1–S6
Pritchard CC, Kroh E, Wood B et al (2012) Blood cell origin of circulating microRNAs: a cautionary note for cancer biomarker studies. Cancer Prev Res (Phila) 5:492–497
Bargaje R, Hariharan M, Scaria V et al (2010) Consensus miRNA expression profiles derived from interplatform normalization of microarray data. RNA 16:16–25
Chang KH, Mestdagh P, Vandesompele J et al (2010) MicroRNA expression profiling to identify and validate reference genes for relative quantification in colorectal cancer. BMC Cancer 10:173
Liang Y, Ridzon D, Wong L et al (2007) Characterization of microRNA expression profiles in normal human tissues. BMC Genomics 8:166
Peltier HJ, Latham GJ (2008) Normalization of microRNA expression levels in quantitative RT-PCR assays: identification of suitable reference RNA targets in normal and cancerous human solid tissues. RNA 14:844–852
Song J, Bai Z, Han W et al (2012) Identification of suitable reference genes for qPCR analysis of serum microRNA in gastric cancer patients. Dig Dis Sci 57:897–904
D’Haene B, Mestdagh P, Hellemans J et al (2012) miRNA expression profiling: from reference genes to global mean normalization. Methods Mol Biol 822:261–272
Wu F, Zhang S, Dassopoulos T et al (2010) Identification of microRNAs associated with ileal and colonic Crohn’s disease. Inflamm Bowel Dis 16:1729–1738
Li Y, Kowdley KV (2012) Method for microRNA isolation from clinical serum samples. Anal Biochem 431:69–75
Mestdagh P, Van Vlierberghe P, De Weer A et al (2009) A novel and universal method for microRNA RT-qPCR data normalization. Genome Biol 10:R64
Acknowledgments
M.C.C. was supported by a postdoctoral fellowship from Fundação para a Ciência e Tecnologia, Portugal (Ref. SFRH/BPD/ 65131/2009).
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Costa, M.C., Leitão, A.L., Enguita, F.J. (2014). MicroRNA Profiling in Plasma or Serum Using Quantitative RT-PCR. In: Alvarez, M., Nourbakhsh, M. (eds) RNA Mapping. Methods in Molecular Biology, vol 1182. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1062-5_11
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DOI: https://doi.org/10.1007/978-1-4939-1062-5_11
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