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Systematic Evaluation of Medium-Throughput mRNA Abundance Platforms

ABSTRACT: Profiling of mRNA abundances with high-throughput platforms such as microarrays and RNA-Seq has become an important tool in both basic and biomedical research. However these platforms remain prone to systematic errors, and have challenges in clinical and industrial application. As a result it is standard practice to validate a subset of key results using alternate technologies. Similarly, clinical and industrial applications typically involve transitions from high-throughput discovery platform to medium-throughput validation ones. These medium-throughput validation platforms have high technical reproducibility and reduced sample input needs, and low sensitivity to sample-quality (e.g. for processing FFPE specimens). Unfortunately, while medium-throughput platforms have proliferated, there are no comprehensive comparisons of them. Here we present ABI's OpenArray and qPCR systems.

ORGANISM(S): Rattus norvegicus

PROVIDER: GSE41388 | GEO | 2012/10/18

SECONDARY ACCESSION(S): PRJNA177846

REPOSITORIES: GEO

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Project description:The study of gene expression as an intermediate phenotype in associating genotype to disease is growing rapidly. The success of this approach depends heavily on the availability of accurate, high throughput platforms for assaying expression levels of genome-wide gene sets. We evaluated a new platform for gene expression, Phalanx Human OneArray (HOA), in 90 HapMap cell lines (CEU panel) for which Illumina BeadChip data were available (Stranger et al. 2007, PMID: 17873874). We found high reproducibility between technical HOA replicates (R = 0.99) and strong inter-platform correlation based on 2,458 sequence-matched probes (R = 0.87). We next performed an association analysis between gene expression levels and HapMap SNPs in the 60 unrelated CEU parents [SNP data downloaded directly from HapMap project (http://www.hapmap.org/)(HapMap phase II/Rel # 22)]. Among the 4356 genes (HOA array) showing variable expression, 122 (2.8%) had a significant cis association (p-value < 0.001) with at least one SNP. Of the 1751 genes showing variable expression on both platforms, 84 had a significant cis association with 36.9% of the genes (31/84) present in both data sets, 39.3 % (33/84) only in HOA, and 23.8% (20/84) only in Illumina. A subset of these 84 eQTLs overlap or are in strong linkage disequilibrium (LD) with reported complex disease/trait susceptibility loci. Combining the results of the two platforms provides a more comprehensive eQTL database (30% increase), which may help to accelerate our understanding of the molecular basis of quantitative traits and diseases. We quantified gene expression levels in the 90 HapMap cell lines (CEU panel) using Phalanx HOA for which Illumina BeadChip data were available (Stranger et al. 2007). We used the same set of RNA samples employed in Stranger et al 2007 (see GEO Series GSE6536, CEU panel; raw data were re-analyzed, ie, re-normalized, thereby generating new values) which allowed us to undertake a detailed comparison to the Illumina expression platform and profiled the 60 parent samples at Industrial Technology Research Institute (ITRI), Taiwan and the 30 children samples at Wellcome Trust Sanger Institute (WTSI), United Kingdom. We performed two in vitro transcription (IVT) reactions for each RNA sample and three technical replicates per IVT with exception of seven samples (15CP, 33CP, 43CP, 44CP, 48CP, 56CP, 60CP) that had two technical replicates for one of the two IVT reactions. *** This Series represents the Phalanx HOA data. *** File "info.pass.parent.txt" (a description of the relationship between an array and a sample and its replicates) is linked below as a supplementary file.

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Systematic Evaluation of Medium-Throughput mRNA Abundance Platforms

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