%0 Journal Article
%T Characterization of the effect of sample quality on high density oligonucleotide microarray data using progressively degraded rat liver RNA
%A Karol L Thompson
%A P Scott Pine
%A Barry A Rosenzweig
%A Yaron Turpaz
%A Jacques Retief
%J BMC Biotechnology
%D 2007
%I BioMed Central
%R 10.1186/1472-6750-7-57
%X Incubation of tissue at 37¡ãC for several hours had little effect on RNA integrity, but did induce changes in the transcript levels of stress response genes and immune cell markers. In contrast, thawing of tissue led to a rapid loss of RNA integrity. Probe sets identified as most sensitive to RNA degradation tended to be located more than 1000 nucleotides upstream of their transcription termini, similar to the positioning of control probe sets used to assess sample quality on Affymetrix GeneChip£¿ arrays. Samples with RNA integrity numbers less than or equal to 7 showed a significant increase in false positives relative to undegraded liver RNA and a reduction in the detection of true positives among probe sets most sensitive to sample integrity for in silico modeled changes of 1.5-, 2-, and 4-fold.Although moderate levels of RNA degradation are tolerated by microarrays with 3'-biased probe selection designs, in this study we identify a threshold beyond which decreased specificity and sensitivity can be observed that closely correlates with average target length. These results highlight the value of annotating microarray data with metrics that capture important aspects of sample quality.It is recommended that the highest quality RNA be used for genomic analyses. However, in some cases, such as human autopsy samples or paraffin embedded tissues, high quality RNA samples may not be available. It is therefore important to understand how RNA quality affects the interpretation of the results and also how reliable current quality measures are at indicating RNA quality issues. It has been reported that gene expression profiling on Affymetrix GeneChip arrays is relatively tolerant to moderate RNA degradation and to the 5'-truncation that occurs during the two successive rounds of in vitro transcription needed to detect small sample quantities [1-3]. Some samples fall within a "grey zone" of sample quality, where there is some loss of RNA integrity but the samples still pass RN
%U http://www.biomedcentral.com/1472-6750/7/57