title: Transcriptome analysis using RNA-Seq.
abstract: Transcriptome analysis by next-generation sequencing (RNA-seq) allows investigation of a transcriptome at unsurpassed resolution. One major benefit is that RNA-seq is independent of a priori knowledge on the sequence under investigation, thereby also allowing analysis of poorly characterized Plasmodium species. Here we provide a detailed protocol for RNA isolation and fragmentation, ribosomal RNA depletion, and cDNA synthesis that enables the preparation of a sequencing library from 1 to 2 μg of total RNA. Although we focus our discussion on the quantitative measurement of gene expression, this protocol is suited for many applications of RNA-seq and allows analysis of most RNA species.
authors.affiliation: Department of Molecular Biology, Nijmegen Center for Molecular Life Sciences, Nijmegen, The Netherlands.
authors.forename: Wieteke A M
authors.initials: WA
authors.lastname: Hoeijmakers
authors.forename: Richárd
authors.initials: R
authors.lastname: Bártfai
authors.forename: Hendrik G
authors.initials: HG
authors.lastname: Stunnenberg
mesh_terms: D020869:Gene Expression Profiling / Q000379:methods*; D015723:Gene Library; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans; D010961:Plasmodium / Q000235:genetics*; D016053:RNA, Protozoan / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D017423:Sequence Analysis, RNA / Q000379:methods*; D059467:Transcriptome
chemical_list: D016053:RNA, Protozoan