title: Genome-wide screen for miRNA targets using the MISSION target ID library.
abstract: The Target ID Library is designed to assist in discovery and identification of microRNA (miRNA) targets. The Target ID Library is a plasmid-based, genome-wide cDNA library cloned into the 3'UTR downstream from the dual-selection fusion protein, thymidine kinase-zeocin (TKzeo). The first round of selection is for stable transformants, followed with introduction of a miRNA of interest, and finally, selecting for cDNAs containing the miRNA's target. Selected cDNAs are identified by sequencing (see Figure 1-3 for Target ID Library Workflow and details).    To ensure broad coverage of the human transcriptome, Target ID Library cDNAs were generated via oligo-dT priming using a pool of total RNA prepared from multiple human tissues and cell lines. Resulting cDNA range from 0.5 to 4 kb, with an average size of 1.2 kb, and were cloned into the p3Î"TKzeo dual-selection plasmid (see Figure 4 for plasmid map). The gene targets represented in the library can be found on the Sigma-Aldrich webpage. Results from Illumina sequencing (Table 3), show that the library includes 16,922 of the 21,518 unique genes in UCSC RefGene (79%), or 14,000 genes with 10 or more reads (66%).
authors.affiliation: Sigma-Aldrich. Matthew.coussens@sial.com
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Coussens
authors.forename: Kevin
authors.initials: K
authors.lastname: Forbes
authors.forename: Carol
authors.initials: C
authors.lastname: Kreader
authors.forename: Jack
authors.initials: J
authors.lastname: Sago
authors.forename: Carrie
authors.initials: C
authors.lastname: Cupp
authors.forename: John
authors.initials: J
authors.lastname: Swarthout
mesh_terms: D020413:3' Untranslated Regions; D002460:Cell Line; D015723:Gene Library; D015894:Genome, Human; D006801:Humans; D035683:MicroRNAs / Q000235:genetics*; D059467:Transcriptome; D014162:Transfection / Q000379:methods
chemical_list: D020413:3' Untranslated Regions; D035683:MicroRNAs