Identification and Correction of Amplification Protocol Bias in Microarray Studies

mRNA abundance microarrays are the most widely used high-throughput technology for functional genomics. But despite this extensive characterization, and despite their wide-spread adoption in discovery research, the uptake of microarrays as clinical diagnostic tools remains relatively limited due to sample degradation. Recently, the NuGEN WT-Ovation Pico kit was developed to resolve this problem. Here, we comprehensively evaluate its utility on two separate biological problems. First, we evaluate its effect on 12 malignant and 2 normal cervix samples. Second, we evaluate its effect on 2 normal kidney samples. In each case, we directly compare aliquots of the same RNA sample that are prepared by standard and low-input/low-RIN protocols to address three key questions. First, does the use of a low-input/degraded-sample protocol alter transcriptional profiles either globally or of specific genes? Second, do these changes alter biological conclusions drawn from the dataset? Third, are these changes biased towards specific classes of genes, either in terms of their sequence characteristics (e.g. length, GC-content) or their biological function?

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