title: Comprehensive identification of miRNA target sites in live animals.
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.
authors: Department of Biology, University of California, La Jolla, San Diego, CA, USA. dzisoulis@ucsd.edu
authors: Dimitrios G
authors: DG
authors: Zisoulis
authors: Gene W
authors: GW
authors: Yeo
authors: Amy E
authors: AE
authors: Pasquinelli
mesh_terms: D000818:Animals; D017173:Caenorhabditis elegans / Q000235:genetics*; D029742:Caenorhabditis elegans Proteins / Q000235:genetics*; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D047468:Immunoprecipitation / Q000379:methods*; D035683:MicroRNAs / Q000235:genetics*; D012333:RNA, Messenger / Q000032:analysis* / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000235:genetics*
chemical_list: C558782:ALG-1 protein, C elegans; D029742:Caenorhabditis elegans Proteins; D035683:MicroRNAs; D012333:RNA, Messenger; D016601:RNA-Binding Proteins