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This article is part of the supplement: EADGENE and SABRE Post-analyses Workshop

Open Access Research

OligoRAP – an Oligo Re-Annotation Pipeline to improve annotation and estimate target specificity

Pieter BT Neerincx1, Han Rauwerda2, Haisheng Nie3, Martien AM Groenen3, Timo M Breit2 and Jack AM Leunissen1*

Author Affiliations

1 Laboratory of Bioinformatics, Wageningen University and Research centre (WUR), P.O. Box 569, 6700 AN Wageningen, The Netherlands

2 Micro array Department (MAD), Swammerdam Institute for Life Sciences, University of Amsterdam (UvA), Kruislaan 318, 1098 SM Amsterdam, The Netherlands

3 Animal Breeding and Genomics Centre, Wageningen University and Research centre (WUR), P.O. Box 338, 6700 AH, Wageningen, The Netherlands

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BMC Proceedings 2009, 3(Suppl 4):S4  doi:10.1186/1753-6561-3-S4-S4

Published: 16 July 2009

Abstract

Background

High throughput gene expression studies using oligonucleotide microarrays depend on the specificity of each oligonucleotide (oligo or probe) for its target gene. However, target specific probes can only be designed when a reference genome of the species at hand were completely sequenced, when this genome were completely annotated and when the genetic variation of the sampled individuals were completely known. Unfortunately there is not a single species for which such a complete data set is available. Therefore, it is important that probe annotation can be updated frequently for optimal interpretation of microarray experiments.

Results

In this paper we present OligoRAP, a pipeline to automatically update the annotation of oligo libraries and estimate oligo target specificity. OligoRAP uses a reference genome assembly with Ensembl and Entrez Gene annotation supplemented with a set of unmapped transcripts derived from RefSeq and UniGene to handle assembly gaps. OligoRAP produces alignments of each oligo with the reference assembly as well as with unmapped transcripts. These alignments are re-mapped to the annotation sources, which results in a concise, as complete as possible and up-to-date annotation of the oligo library. The building blocks of this pipeline are BioMoby web services creating a highly modular and distributed system with a robust, remote programmatic interface.

OligoRAP was used to update the annotation for a subset of 791 oligos from the ARK-Genomics 20 K chicken array, which were selected as starting material for the oligo annotation session of the EADGENE/SABRE Post-analysis workshop. Based on the updated annotation about one third of these oligos is problematic with regard to target specificity. In addition, the accession numbers or ids the oligos were originally designed for no longer exist in the updated annotation for almost half of the oligos.

Conclusion

As microarrays are designed on incomplete data, it is important to update probe annotation and check target specificity regularly. OligoRAP provides both and due to its design based on BioMoby web services it can easily be embedded as an oligo annotation engine in customised applications for microarray data analysis. The dramatic difference in updated annotation and target specificity for the ARK-Genomics 20 K chicken array as compared to the original data emphasises the need for regular updates.