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      Estimation of data-specific constitutive exons with RNA-Seq data

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      BMC Bioinformatics
      BioMed Central

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          Abstract

          Background

          RNA-Seq has the potential to answer many diverse and interesting questions about the inner workings of cells. Estimating changes in the overall transcription of a gene is not straightforward. Changes in overall gene transcription can easily be confounded with changes in exon usage which alter the lengths of transcripts produced by a gene. Measuring the expression of constitutive exons— exons which are consistently conserved after splicing— offers an unbiased estimation of the overall transcription of a gene.

          Results

          We propose a clustering-based method, exClust, for estimating the exons that are consistently conserved after splicing in a given data set. These are considered as the exons which are “constitutive” in this data. The method utilises information from both annotation and the dataset of interest. The method is implemented in an openly available R function package, sydSeq.

          Conclusion

          When used on two real datasets exClust includes more than three times as many reads as the standard UI method, and improves concordance with qRT-PCR data. When compared to other methods, our method is shown to produce robust estimates of overall gene transcription.

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          Most cited references14

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          Ensembl 2009

          The Ensembl project (http://www.ensembl.org) is a comprehensive genome information system featuring an integrated set of genome annotation, databases, and other information for chordate, selected model organism and disease vector genomes. As of release 51 (November 2008), Ensembl fully supports 45 species, and three additional species have preliminary support. New species in the past year include orangutan and six additional low coverage mammalian genomes. Major additions and improvements to Ensembl since our previous report include a major redesign of our website; generation of multiple genome alignments and ancestral sequences using the new Enredo-Pecan-Ortheus pipeline and development of our software infrastructure, particularly to support the Ensembl Genomes project (http://www.ensemblgenomes.org/).
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            From RNA-seq reads to differential expression results

            Many methods and tools are available for preprocessing high-throughput RNA sequencing data and detecting differential expression.
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              Transcript length bias in RNA-seq data confounds systems biology

              Background Several recent studies have demonstrated the effectiveness of deep sequencing for transcriptome analysis (RNA-seq) in mammals. As RNA-seq becomes more affordable, whole genome transcriptional profiling is likely to become the platform of choice for species with good genomic sequences. As yet, a rigorous analysis methodology has not been developed and we are still in the stages of exploring the features of the data. Results We investigated the effect of transcript length bias in RNA-seq data using three different published data sets. For standard analyses using aggregated tag counts for each gene, the ability to call differentially expressed genes between samples is strongly associated with the length of the transcript. Conclusion Transcript length bias for calling differentially expressed genes is a general feature of current protocols for RNA-seq technology. This has implications for the ranking of differentially expressed genes, and in particular may introduce bias in gene set testing for pathway analysis and other multi-gene systems biology analyses. Reviewers This article was reviewed by Rohan Williams (nominated by Gavin Huttley), Nicole Cloonan (nominated by Mark Ragan) and James Bullard (nominated by Sandrine Dudoit).
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                Author and article information

                Contributors
                Journal
                BMC Bioinformatics
                BMC Bioinformatics
                BMC Bioinformatics
                BioMed Central
                1471-2105
                2013
                29 January 2013
                : 14
                : 31
                Affiliations
                [1 ]School of Mathematics and Statistics, University of Sydney, Sydney NSW 2006, Australia
                [2 ]CSIRO Mathematical & Information Sciences, Private Bag 33, Clayton South 3168, Australia
                Article
                1471-2105-14-31
                10.1186/1471-2105-14-31
                3656776
                23360225
                81da0b22-1634-4645-be2f-992377174828
                Copyright © 2013 Patrick et al.; licensee BioMed Central Ltd.

                This is an Open Access article distributed under the terms of the Creative Commons Attribution License ( http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

                History
                : 25 April 2012
                : 13 January 2013
                Categories
                Methodology Article

                Bioinformatics & Computational biology
                Bioinformatics & Computational biology

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