Abstract
MicroRNAs (miRNAs) are small RNA molecules that posttranscriptionally regulate the expression of protein-coding genes. The mature miRNAs are loaded into Argonaute-containing protein complexes (miRISC, miRNA Induced S ilencing Complex), and guide these complexes to the 3′ UTR of targeted mRNA transcripts via base-pairing interactions. However, the imperfect complementarity that characterizes the interactions between animal miRNAs and target sites complicates the identification of direct target genes. We developed a biochemical method to identify on a large scale the target sequences recognized by miRISC in vivo. The mRNA sites bound by miRISC are stabilized by cross-linking and isolated by immunoprecipitation of Argonaute-containing complexes. The bound RNA molecules are trimmed to the regions protected by Argonaute, subjected to a series of isolation and linker ligation steps and identified by high-throughput sequencing methods.
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© 2011 Humana Press
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Zisoulis, D.G., Yeo, G.W., Pasquinelli, A.E. (2011). Comprehensive Identification of miRNA Target Sites in Live Animals. In: Dalmay, T. (eds) MicroRNAs in Development. Methods in Molecular Biology, vol 732. Humana Press. https://doi.org/10.1007/978-1-61779-083-6_13
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DOI: https://doi.org/10.1007/978-1-61779-083-6_13
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