Transcriptome sequencing of the Microarray Quality Control (MAQC) RNA reference samples using next generation sequencing.
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ABSTRACT: Background
Transcriptome sequencing using next-generation sequencing platforms will soon be competing with DNA microarray technologies for global gene expression analysis. As a preliminary evaluation of these promising technologies, we performed deep sequencing of cDNA synthesized from the Microarray Quality Control (MAQC) reference RNA samples using Roche's 454 Genome Sequencer FLX.Results
We generated more that 3.6 million sequence reads of average length 250 bp for the MAQC A and B samples and introduced a data analysis pipeline for translating cDNA read counts into gene expression levels. Using BLAST, 90% of the reads mapped to the human genome and 64% of the reads mapped to the RefSeq database of well annotated genes with e-values ConclusionUsing the MAQC metrics for evaluating the performance of gene expression platforms, the ExpressSeq results for gene expression levels showed excellent reproducibility, sensitivity, and specificity that improved systematically with increasing shotgun sequencing depth, and quantitative accuracy that was comparable to DNA microarrays and QRTPCR. In addition, a careful mapping of the reads to the genome using the AceView alignment programs shed new light on the complexity of the human transcriptome including the discovery of thousands of new splice variants.
SUBMITTER: Mane SP
PROVIDER: S-EPMC2707382 | biostudies-literature | 2009 Jun
REPOSITORIES: biostudies-literature
Publications
Transcriptome sequencing of the Microarray Quality Control (MAQC) RNA reference samples using next generation sequencing.
Mane Shrinivasrao P SP Evans Clive C Cooper Kristal L KL Crasta Oswald R OR Folkerts Otto O Hutchison Stephen K SK Harkins Timothy T TT Thierry-Mieg Danielle D Thierry-Mieg Jean J Jensen Roderick V RV
BMC genomics 20090612
<h4>Background</h4>Transcriptome sequencing using next-generation sequencing platforms will soon be competing with DNA microarray technologies for global gene expression analysis. As a preliminary evaluation of these promising technologies, we performed deep sequencing of cDNA synthesized from the Microarray Quality Control (MAQC) reference RNA samples using Roche's 454 Genome Sequencer FLX.<h4>Results</h4>We generated more that 3.6 million sequence reads of average length 250 bp for the MAQC A ...[more]
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