title: High-throughput multiplex sequencing of miRNA.
abstract: Next-generation sequencing offers many advantages over other methods of microRNA (miRNA) expression profiling, such as sample throughput and the capability to discover novel miRNAs. As the sequencing depth of current sequencing platforms exceeds what is necessary to quantify miRNAs, multiplexing several samples in one sequencing run offers a significant cost advantage. Although previous studies have achieved this goal by adding bar codes to miRNA libraries at the ligation step, this was recently shown to introduce significant bias into the miRNA expression data. This bias can be avoided, however, by bar coding the miRNA libraries at the PCR step instead. Here, we describe a user-friendly PCR bar-coding method of preparing multiplexed microRNA libraries for Illumina-based sequencing. The method also prevents the production of adapter dimers and can be completed in one day.
authors.affiliation: Department of Genetics, Harvard Medical School, Boston, Massachusetts.|Wyss Institute for Biologically Inspired Engineering, Boston, Massachusetts.|Ragon Institute of MGH, MIT, and Harvard, Charlestown, Massachusetts.
authors.forename: Francois
authors.initials: F
authors.lastname: Vigneault
authors.affiliation: Wyss Institute for Biologically Inspired Engineering, Boston, Massachusetts.|Department of Molecular and Cellular Biology, Harvard University, Cambridge, Massachusetts.
authors.forename: Dmitry
authors.initials: D
authors.lastname: Ter-Ovanesyan
authors.affiliation: Department of Neurobiology, George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv, Israel.|Raymond and Beverly Sackler School of Physics and Astronomy, Tel Aviv University, Tel Aviv, Israel.
authors.forename: Shahar
authors.initials: S
authors.lastname: Alon
authors.affiliation: Department of Genetics, Harvard Medical School, Boston, Massachusetts.
authors.forename: Seda
authors.initials: S
authors.lastname: Eminaga
authors.affiliation: Department of Genetics, Harvard Medical School, Boston, Massachusetts.
authors.forename: Danos
authors.initials: D
authors.lastname: C Christodoulou
authors.affiliation: Department of Genetics, Harvard Medical School, Boston, Massachusetts.
authors.forename: J G
authors.initials: JG
authors.lastname: Seidman
authors.affiliation: Department of Neurobiology, George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv, Israel.|Raymond and Beverly Sackler School of Physics and Astronomy, Tel Aviv University, Tel Aviv, Israel.
authors.forename: Eli
authors.initials: E
authors.lastname: Eisenberg
authors.affiliation: Department of Genetics, Harvard Medical School, Boston, Massachusetts.|Wyss Institute for Biologically Inspired Engineering, Boston, Massachusetts.
authors.forename: George
authors.initials: G
authors.lastname: M Church
mesh_terms: D015723:Gene Library; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans; D035683:MicroRNAs / Q000737:chemistry*; D017423:Sequence Analysis, RNA / Q000379:methods*
chemical_list: D035683:MicroRNAs