Transcript-based redefinition of grouped oligonucleotide probe sets using AceView: high-resolution annotation for microarrays.
BACKGROUND
Extracting biological information from high-density Affymetrix arrays is a multi-step process that begins with the accurate annotation of microarray probes. Shortfalls in the original Affymetrix probe annotation have been described; however, few studies have provided rigorous solutions for routine data analysis.


RESULTS
Using AceView, a comprehensive human transcript database, we have reannotated the probes by matching them to RNA transcripts instead of genes. Based on this transcript-level annotation, a new probe set definition was created in which every probe in a probe set maps to a common set of AceView gene transcripts. In addition, using artificial data sets we identified that a minimal probe set size of 4 is necessary for reliable statistical summarization. We further demonstrate that applying the new probe set definition can detect specific transcript variants contributing to differential expression and it also improves cross-platform concordance.


CONCLUSIONS
We conclude that our transcript-level reannotation and redefinition of probe sets complement the original Affymetrix design. Redefinitions introduce probe sets whose sizes may not support reliable statistical summarization; therefore, we advocate using our transcript-level mapping redefinition in a secondary analysis step rather than as a replacement. Knowing which specific transcripts are differentially expressed is important to properly design probe/primer pairs for validation purposes. For convenience, we have created custom chip-description-files (CDFs) and annotation files for our new probe set definitions that are compatible with Bioconductor, Affymetrix Expression Console or third party software.
Lu
Jun
J
Genomics Core Laboratory, National Institute of Diabetes & Digestive & Kidney Diseases, National Institutes of Health, Bethesda, MD 20892 USA. lujun@mail.nih.gov <lujun@mail.nih.gov>
Lee
Joseph C
JC
Salit
Marc L
ML
Cam
Margaret C
MC
D002874:Chromosome Mapping / Q000379:methods*; D030541:Databases, Genetic; D004282:Documentation / Q000379:methods*; D020869:Gene Expression Profiling / Q000379:methods*; D006801:Humans; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D015345:Oligonucleotide Probes / Q000235:genetics*; D012680:Sensitivity and Specificity; D014157:Transcription Factors / Q000235:genetics*
D015345:Oligonucleotide Probes; D014157:Transcription Factors