Abstract
As small noncoding RNAs, microRNAs (miRNAs) regulate diverse biological functions, including physiological and pathological processes. The expression and deregulation of miRNA levels contain rich information with diagnostic and prognostic relevance and can reflect pharmacological responses. The increasing interest in miRNA-related research demands global miRNA expression profiling on large numbers of samples. We describe here a robust protocol that supports high-throughput sample labeling and detection on hundreds of samples simultaneously. This method employs 96-well-based miRNA capturing from total RNA samples and on-site biochemical reactions, coupled with bead-based detection in 96-well format for hundreds of miRNAs per sample. With low-cost, high-throughput, high detection specificity, and flexibility to profile both small and large numbers of samples, this protocol can be adapted in a wide range of laboratory settings.
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References
Bartel DP (2009) MicroRNAs: target recognition and regulatory functions. Cell 136:215–233
Ambros V (2004) The functions of animal microRNAs. Nature 431:350–355
Garzon R, Croce CM (2008) MicroRNAs in normal and malignant hematopoiesis. Curr Opin Hematol 15:352–358
Farazi TA, Spitzer JI, Morozov P et al (2011) miRNAs in human cancer. J Pathol 223:102–115
Lu J, Getz G, Miska EA et al (2005) MicroRNA expression profiles classify human cancers. Nature 435:834–838
Wang Y, Juranek S, Li H et al (2008) Structure of an argonaute silencing complex with a seed-containing guide DNA and target RNA duplex. Nature 456:921–926
D’Andrade PN, Fulmer-Smentek S (2012) Agilent microRNA microarray profiling system. Methods Mol Biol 822:85–102
Lu J, Guo S, Ebert BL et al (2008) MicroRNA-mediated control of cell fate in megakaryocyte-erythrocyte progenitors. Dev Cell 14:843–853
Li Z, Lu J, Sun M et al (2008) Distinct microRNA expression profiles in acute myeloid leukemia with common translocations. Proc Natl Acad Sci U S A 105:15535–15540
Olson P, Lu J, Zhang H et al (2009) MicroRNA dynamics in the stages of tumorigenesis correlate with hallmark capabilities of cancer. Genes Dev 23:2152–2165
Mi S, Lu J, Sun M et al (2007) MicroRNA expression signatures accurately discriminate acute lymphoblastic leukemia from acute myeloid leukemia. Proc Natl Acad Sci U S A 104:19971–19976
Adams BD, Guo S, Bai H et al (2012) An in vivo functional screen uncovers miR-150-mediated regulation of hematopoietic injury response. Cell Rep 2:1048–1060
Guo S, Bai H, Megyola CM et al (2012) Complex oncogene dependence in microRNA-125a-induced myeloproliferative neoplasms. Proc Natl Acad Sci U S A 109:16636–16641
Guo S, Lu J, Schlanger R et al (2010) MicroRNA miR-125a controls hematopoietic stem cell number. Proc Natl Acad Sci U S A 107:14229–14234
Hafner M, Landgraf P, Ludwig J et al (2008) Identification of microRNAs and other small regulatory RNAs using cDNA library sequencing. Methods 44:3–12
Vigneault F, Sismour AM, Church GM (2008) Efficient microRNA capture and bar-coding via enzymatic oligonucleotide adenylation. Nat Methods 5:777–779
Acknowledgments
This work was supported in part by NIH grants R01CA149109 and R01GM099811 and Connecticut Stem Cell Research Fund (09SCBYALE27). We thank Eric Miska, Ezequiel Alvarez-Saavedra, Justin Lamb, David Peck, Hao Zhang, and Judy Wang for help during the process of establishing this protocol.
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Guo, Y., Mastriano, S., Lu, J. (2014). A High-Throughput MicroRNA Expression Profiling System. In: Wajapeyee, N. (eds) Cancer Genomics and Proteomics. Methods in Molecular Biology, vol 1176. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-0992-6_4
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DOI: https://doi.org/10.1007/978-1-4939-0992-6_4
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Publisher Name: Humana Press, New York, NY
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Online ISBN: 978-1-4939-0992-6
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