A comparison of massively parallel nucleotide sequencing with oligonucleotide microarrays for global transcription profiling.
AffiliationApplied Computational Biology and Bioinformatics, Cancer Research UK, Paterson Institute for Cancer Research, The University of Manchester, Wilmslow Road, Manchester M20 4BX UK.
MetadataShow full item record
AbstractBACKGROUND: RNA-Seq exploits the rapid generation of gigabases of sequence data by Massively Parallel Nucleotide Sequencing, allowing for the mapping and digital quantification of whole transcriptomes. Whilst previous comparisons between RNA-Seq and microarrays have been performed at the level of gene expression, in this study we adopt a more fine-grained approach. Using RNA samples from a normal human breast epithelial cell line (MCF-10a) and a breast cancer cell line (MCF-7), we present a comprehensive comparison between RNA-Seq data generated on the Applied Biosystems SOLiD platform and data from Affymetrix Exon 1.0ST arrays. The use of Exon arrays makes it possible to assess the performance of RNA-Seq in two key areas: detection of expression at the granularity of individual exons, and discovery of transcription outside annotated loci. RESULTS: We found a high degree of correspondence between the two platforms in terms of exon-level fold changes and detection. For example, over 80% of exons detected as expressed in RNA-Seq were also detected on the Exon array, and 91% of exons flagged as changing from Absent to Present on at least one platform had fold-changes in the same direction. The greatest detection correspondence was seen when the read count threshold at which to flag exons Absent in the SOLiD data was set to t<1 suggesting that the background error rate is extremely low in RNA-Seq. We also found RNA-Seq more sensitive to detecting differentially expressed exons than the Exon array, reflecting the wider dynamic range achievable on the SOLiD platform. In addition, we find significant evidence of novel protein coding regions outside known exons, 93% of which map to Exon array probesets, and are able to infer the presence of thousands of novel transcripts through the detection of previously unreported exon-exon junctions. CONCLUSIONS: By focusing on exon-level expression, we present the most fine-grained comparison between RNA-Seq and microarrays to date. Overall, our study demonstrates that data from a SOLiD RNA-Seq experiment are sufficient to generate results comparable to those produced from Affymetrix Exon arrays, even using only a single replicate from each platform, and when presented with a large genome.
CitationA comparison of massively parallel nucleotide sequencing with oligonucleotide microarrays for global transcription profiling. 2010, 11:282 BMC Genomics
- Gene expression and isoform variation analysis using Affymetrix Exon Arrays.
- Authors: Bemmo A, Benovoy D, Kwan T, Gaffney DJ, Jensen RV, Majewski J
- Issue date: 2008 Nov 7
- A systematic comparison and evaluation of high density exon arrays and RNA-seq technology used to unravel the peripheral blood transcriptome of sickle cell disease.
- Authors: Raghavachari N, Barb J, Yang Y, Liu P, Woodhouse K, Levy D, O'Donnell CJ, Munson PJ, Kato GJ
- Issue date: 2012 Jun 29
- Comparison of Affymetrix Gene Array with the Exon Array shows potential application for detection of transcript isoform variation.
- Authors: Ha KCh, Coulombe-Huntington J, Majewski J
- Issue date: 2009 Nov 12
- A comparison of RNA-seq and exon arrays for whole genome transcription profiling of the L5 spinal nerve transection model of neuropathic pain in the rat.
- Authors: Perkins JR, Antunes-Martins A, Calvo M, Grist J, Rust W, Schmid R, Hildebrandt T, Kohl M, Orengo C, McMahon SB, Bennett DL
- Issue date: 2014 Jan 28
- RNA sequencing and transcriptome arrays analyses show opposing results for alternative splicing in patient derived samples.
- Authors: Nazarov PV, Muller A, Kaoma T, Nicot N, Maximo C, Birembaut P, Tran NL, Dittmar G, Vallar L
- Issue date: 2017 Jun 6