title: De novo transcript sequence reconstruction from RNA-seq using the Trinity platform for reference generation and analysis.
abstract: De novo assembly of RNA-seq data enables researchers to study transcriptomes without the need for a genome sequence; this approach can be usefully applied, for instance, in research on 'non-model organisms' of ecological and evolutionary importance, cancer samples or the microbiome. In this protocol we describe the use of the Trinity platform for de novo transcriptome assembly from RNA-seq data in non-model organisms. We also present Trinity-supported companion utilities for downstream applications, including RSEM for transcript abundance estimation, R/Bioconductor packages for identifying differentially expressed transcripts across samples and approaches to identify protein-coding genes. In the procedure, we provide a workflow for genome-independent transcriptome analysis leveraging the Trinity platform. The software, documentation and demonstrations are freely available from http://trinityrnaseq.sourceforge.net. The run time of this protocol is highly dependent on the size and complexity of data to be analyzed. The example data set analyzed in the procedure detailed herein can be processed in less than 5 h.
authors.affiliation: Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA, 02142, USA.
authors.forename: Brian J
authors.initials: BJ
authors.lastname: Haas
authors.affiliation: CSIRO Ecosystem Sciences, Black Mountain Labs, Canberra, ACT 2601, Australia.
authors.forename: Alexie
authors.initials: A
authors.lastname: Papanicolaou
authors.affiliation: Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA, 02142, USA.|The Selim and Rachel Benin School of Computer Science, The Hebrew University of Jerusalem, Jerusalem 91904, Israel.
authors.forename: Moran
authors.initials: M
authors.lastname: Yassour
authors.affiliation: Science for Life Laboratory, Department of Medical Biochemistry and Microbiology, Uppsala University, Uppsala, Sweden.
authors.forename: Manfred
authors.initials: M
authors.lastname: Grabherr
authors.affiliation: Pittsburgh Supercomputing Center, Carnegie Mellon University, Pittsburgh, PA, 15213, USA.
authors.forename: Philip D
authors.initials: PD
authors.lastname: Blood
authors.affiliation: CSIRO Information Management & Technology, 306 Carmody Rd, St Lucia QLD 4067, Australia.
authors.forename: Joshua
authors.initials: J
authors.lastname: Bowden
authors.affiliation: Department of Microbiology and Molecular Genetics, Oklahoma State University, USA.
authors.forename: Matthew Brian
authors.initials: MB
authors.lastname: Couger
authors.affiliation: Genomics Research Centre, Griffith University, Gold Coast Campus, Qld 4222, Australia.
authors.forename: David
authors.initials: D
authors.lastname: Eccles
authors.affiliation: Department of Computer Sciences, University of Wisconsin, Madison, WI, 53706, USA.
authors.forename: Bo
authors.initials: B
authors.lastname: Li
authors.affiliation: Technische Universität Dresden, Dresden, Saxony 01062, Germany.
authors.forename: Matthias
authors.initials: M
authors.lastname: Lieber
authors.affiliation: University of California, Berkeley and California Institute for Quantitative Biosciences Berkeley, CA 94720, USA.
authors.forename: Matthew D
authors.initials: MD
authors.lastname: MacManes
authors.affiliation: CSIRO Ecosystem Sciences, Black Mountain Labs, Canberra, ACT 2601, Australia.
authors.forename: Michael
authors.initials: M
authors.lastname: Ott
authors.affiliation: Institute for Genome Sciences, Baltimore, MD, 21201, USA.
authors.forename: Joshua
authors.initials: J
authors.lastname: Orvis
authors.affiliation: Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA, 02142, USA.|Department of Plant Systems Biology, VIB, Department of Plant Biotechnology and Bioinformatics, Ghent University, Ghent B-9052, Belgium.
authors.forename: Nathalie
authors.initials: N
authors.lastname: Pochet
authors.affiliation: Parco Tecnologico Padano, Loc. Cascina Codazza, 26900 Lodi, Italy.
authors.forename: Francesco
authors.initials: F
authors.lastname: Strozzi
authors.affiliation: Corn Insects and Crop Genetics Research Unit, United States Department of Agriculture--Agricultural Research Service, Ames, IA 50011, USA.
authors.forename: Nathan
authors.initials: N
authors.lastname: Weeks
authors.affiliation: Genomics facility, Purdue University, West Lafayette, IN, 47907, USA.
authors.forename: Rick
authors.initials: R
authors.lastname: Westerman
authors.affiliation: GWT-TUD GmbH, Blasewitzer Strasse 43, Dresden, Saxony 01307, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: William
authors.affiliation: Department of Computer Sciences, University of Wisconsin, Madison, WI, 53706, USA.|Department of Biostatistics and Medical Informatics, University of Wisconsin, Madison, WI 53706, USA.
authors.forename: Colin N
authors.initials: CN
authors.lastname: Dewey
authors.affiliation: Indiana University, 2709 East 10th Street, Bloomington, IN 47408,USA.
authors.forename: Robert
authors.initials: R
authors.lastname: Henschel
authors.affiliation: Indiana University, 2709 East 10th Street, Bloomington, IN 47408,USA.
authors.forename: Richard D
authors.initials: RD
authors.lastname: LeDuc
authors.affiliation: The Selim and Rachel Benin School of Computer Science, The Hebrew University of Jerusalem, Jerusalem 91904, Israel.
authors.forename: Nir
authors.initials: N
authors.lastname: Friedman
authors.affiliation: Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA, 02142, USA.|Howard Hughes Medical Institute, Department of Biology, Massachusetts Institute of Technology, Cambridge, MA, 02140.
authors.forename: Aviv
authors.initials: A
authors.lastname: Regev
mesh_terms: D001483:Base Sequence; D020869:Gene Expression Profiling / Q000379:methods*; D012313:RNA / Q000737:chemistry*; D012568:Schizosaccharomyces / Q000235:genetics; D029702:Schizosaccharomyces pombe Proteins / Q000737:chemistry / Q000235:genetics; D017423:Sequence Analysis, RNA / Q000379:methods; D012984:Software; D059467:Transcriptome
chemical_list: D029702:Schizosaccharomyces pombe Proteins; D012313:RNA