title: Hyb: a bioinformatics pipeline for the analysis of CLASH (crosslinking, ligation and sequencing of hybrids) data.
abstract: Associations between proteins and RNA-RNA duplexes are important in post-transcriptional regulation of gene expression. The CLASH (Cross-linking, Ligation and Sequencing of Hybrids) technique captures RNA-RNA interactions by physically joining two RNA molecules associated with a protein complex into a single chimeric RNA molecule. These events are relatively rare and considerable effort is needed to detect a small number of chimeric sequences amongst millions of non-chimeric cDNA reads resulting from a CLASH experiment. We present the "hyb" bioinformatics pipeline, which we developed to analyse high-throughput cDNA sequencing data from CLASH experiments. Although primarily designed for use with AGO CLASH data, hyb can also be used for the detection and annotation of chimeric reads in other high-throughput sequencing datasets. We examined the sensitivity and specificity of chimera detection in a test dataset using the BLAST, BLAST+, BLAT, pBLAT and Bowtie2 read alignment programs. We obtained the most reliable results in the shortest time using a combination of preprocessing with Flexbar and subsequent read-mapping using Bowtie2. The "hyb" software is distributed under the GNU GPL (General Public License) and can be downloaded from https://github.com/gkudla/hyb.
authors.affiliation: Wellcome Trust Centre for Cell Biology, University of Edinburgh, Edinburgh, Scotland, United Kingdom; Institute of Biological and Environmental Sciences, University of Aberdeen, Aberdeen, Scotland, United Kingdom.
authors.forename: Anthony J
authors.initials: AJ
authors.lastname: Travis
authors.affiliation: MRC Human Genetics Unit, Institute of Genetics and Molecular Medicine, University of Edinburgh, Edinburgh, Scotland, United Kingdom.
authors.forename: Jonathan
authors.initials: J
authors.lastname: Moody
authors.affiliation: Wellcome Trust Centre for Cell Biology, University of Edinburgh, Edinburgh, Scotland, United Kingdom.
authors.forename: Aleksandra
authors.initials: A
authors.lastname: Helwak
authors.affiliation: Wellcome Trust Centre for Cell Biology, University of Edinburgh, Edinburgh, Scotland, United Kingdom.
authors.forename: David
authors.initials: D
authors.lastname: Tollervey
authors.affiliation: MRC Human Genetics Unit, Institute of Genetics and Molecular Medicine, University of Edinburgh, Edinburgh, Scotland, United Kingdom. Electronic address: gkudla@gmail.com.
authors.forename: Grzegorz
authors.initials: G
authors.lastname: Kudla
mesh_terms: D020029:Base Pairing; D019295:Computational Biology / Q000379:methods*; D018076:DNA, Complementary / Q000737:chemistry* / Q000235:genetics; D015894:Genome, Human; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D059370:RNA Folding; D012330:RNA, Double-Stranded / Q000737:chemistry* / Q000235:genetics; D012680:Sensitivity and Specificity; D016415:Sequence Alignment; D012984:Software
chemical_list: D018076:DNA, Complementary; D012330:RNA, Double-Stranded
keywords: Bioinformatics; CLASH; High-throughput sequencing; RNA–RNA interactions