RNA-SeQC: RNA-Seq metrics for quality control and process optimization
ABSTRACT Summary: RNA-seq, the application of next-generation sequencing to RNA, provides transcriptome-wide characterization of cellular activity. Assessment of sequencing performance and library quality is critical to the interpretation of RNA-seq data, yet few tools exist to address this issue. We introduce RNA-SeQC, a program which provides key measures of data quality. These metrics include yield, alignment and duplication rates; GC bias, rRNA content, regions of alignment (exon, intron and intragenic), continuity of coverage, 3′/5′ bias and count of detectable transcripts, among others. The software provides multi-sample evaluation of library construction protocols, input materials and other experimental parameters. The modularity of the software enables pipeline integration and the routine monitoring of key measures of data quality such as the number of alignable reads, duplication rates and rRNA contamination. RNA-SeQC allows investigators to make informed decisions about sample inclusion in downstream analysis. In summary, RNA-SeQC provides quality control measures critical to experiment design, process optimization and downstream computational analysis.Availability and implementation: See www.genepattern.org to run online, or www.broadinstitute.org/rna-seqc/ for a command line tool.Contact:
Supplementary data are available at Bioinformatics online.
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Article: RNA-SeQC: RNA-Seq metrics for quality control and process optimization
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- "0 . 11 ; ( Kim et al . , 2013 ) ) , transcript structure and abundance were estimated using Cufflinks ( ver . 2 . 1 . 1 ; ( Trapnell et al . , 2010 ) ) , and differential expression analysis was performed using Cuffdiff ( ver . 2 . 1 . 1 ; ( Trapnell et al . , 2013 ) ) . Quality control analysis was performed using RNA - SeQC ( ver . 1 . 1 . 7 ; ( DeLuca et al . , 2012 ) ) . The cummeRbund package ( ver . 2 . 4 . 1 ; ( Trapnell et al . , 2012 ) ) for R ( ver . 3 . 0 . 2 ) was used for data visualization . Differen - tial expression analysis was performed for the four donor samples Table 3 Top 40 differentially expressed genes between temporal retina vs . macular retina with q - value < 0 . 001 and abs"
ABSTRACT: We examined gene expression in temporal, macular, and nasal regions of human retina and retinal pigment epithelium (RPE)/choroid using RNA-Seq.•Expression differences between macula and periphery (nasal and temporal regions) in both tissues reflect the distribution of cell types.•Nasal and temporal regions of neural retina are indistinguishable in our analysis.Experimental Eye Research 11/2014; 129. DOI:10.1016/j.exer.2014.11.001
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- "The mapped reads are available as.sam or.bam files for each sample, which needs to be quality controlled because some issues only appear after the mapping/alignment of reads are finished. Running the after-alignment/mapping quality control (e.g. using software such as RNA-SeQC, DeLuca et al., 2012) or Qualimap (http://qualimap.bioinfo. cipf.es/) "
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- "Resource Description References GeNCODe annotates gene-based features including alternatively transcribed variants PMID: 22955987 Harrow et al. (2012) eNCODe Catalogs all functional elements in the genome PMID: 15499007 eNCODe Project Consortium (2004) ensembl Produces genome databases for vertebrates and other eukaryotic species PMID: 11752248 Hubbard et al. (2002) RefSeq Provides reference sequence standards for genomes, transcripts and proteins PMID: 11125071 Pruitt and Maglott (2001) GTex Provides a resource database and tissue bank to investigate relationship between genetic variation and gene expression in human tissues PMID: 23715323 (GTex Consortium 2013) Illumina Human Body Map Provides reference RNa-Seq data in 16 different human tissues PMID: 22496456 asmann et al. (2012) RNa-SeQC Generates quality control (QC) metrics for RNa-Seq data PMID: 22539670 DeLuca et al. (2012) htSeqTools Provides quality assessment and visualization of highthroughput data in the Bioconductor environment PMID: 22199381 Planet et al. (2012) TopHat and Cufflinks aligns reads, identifies splice sites, and performs differential expression analysis of RNa-Seq data PMID: 22383036 Trapnell et al. (2012) DeSeq and edgeR Facilitates differential expression analysis of RNa-Seq data using R and Bioconductor PMID: 23975260 anders et al. (2013) aStalavista extracts and displays alternative splicing events PMID: 17485470 Foissac and Sammeth (2007) eCGene Provides annotation for gene structure, function and expression using alternative splicing PMID: 15608289 Kim et al. (2005) MISO/sashimi_plot Quantifies alternatively spliced genes from RNa-Seq and provides visualization PMID: 21057496 Katz et al. (2010) Integrated Genomics viewer Facilitates visualization of genomics high-throughput data PMID: 21221095 Robinson et al. (2011) ReSCUe-eSe Identifies sequences with exonic splicing enhancer activity PMID: 12114529 Fairbrother et al. (2002) FaS-eSS Identifies sequences with exonic splicing silencer activity PMID: 15607979 wang et al. (2004) expression as eQTLs (Coulombe-Huntington et al. 2009). variation in alternative splicing is highly heritable, with family-based linkage analysis demonstrating that transcript isoforms of a variety of genes undergo Mendelian inheritance and segregation (Kwan et al. 2007). "
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