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Whole-genome microarrays of fission yeast: characteristics, accuracy, reproducibility, and processing of array data

Lyne, R; Burns, G; Mata, J; Penkett, CJ; Rustici, G; Chen, DR; Langford, C; ... Bahler, J; + view all (2003) Whole-genome microarrays of fission yeast: characteristics, accuracy, reproducibility, and processing of array data. BMC GENOMICS , 4 , Article 27. 10.1186/1471-2164-4-27. Green open access

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Abstract

Background: The genome of the fission yeast Schizosaccharomyces pombe has recently been sequenced, setting the stage for the post-genomic era of this increasingly popular model organism. We have built fission yeast microarrays, optimised protocols to improve array performance, and carried out experiments to assess various characteristics of microarrays.Results: We designed PCR primers to amplify specific probes (180-500 bp) for all known and predicted fission yeast genes, which are printed in duplicate onto separate regions of glass slides together with control elements (similar to13,000 spots/slide). Fluorescence signal intensities depended on the size and intragenic position of the array elements, whereas the signal ratios were largely independent of element properties. Only the coding strand is covalently linked to the slides, and our array elements can discriminate transcriptional direction. The microarrays can distinguish sequences with up to 70% identity, above which cross-hybridisation contributes to the signal intensity. We tested the accuracy of signal ratios and measured the reproducibility of array data caused by biological and technical factors. Because the technical variability is lower, it is best to use samples prepared from independent biological experiments to obtain repeated measurements with swapping of fluorochromes to prevent dye bias. We also developed a script that discards unreliable data and performs a normalization to correct spatial artefacts.Conclusions: This paper provides data for several microarray properties that are rarely measured. The results define critical parameters for microarray design and experiments and provide a framework to optimise and interpret array data. Our arrays give reproducible and accurate expression ratios with high sensitivity. The scripts for primer design and initial data processing as well as primer sequences and detailed protocols are available from our website.

Type: Article
Title: Whole-genome microarrays of fission yeast: characteristics, accuracy, reproducibility, and processing of array data
Open access status: An open access version is available from UCL Discovery
DOI: 10.1186/1471-2164-4-27
Publisher version: http://dx.doi.org/10.1186/1471-2164-4-27
Language: English
Additional information: © 2003 Lyne et al; licensee BioMed Central Ltd. This is an Open Access article: verbatim copying and redistribution of this article are permitted in all media for any purpose, provided this notice is preserved along with the article's original URL
Keywords: SCHIZOSACCHAROMYCES-POMBE, DNA MICROARRAY, GENE-EXPRESSION, MEIOSIS, HYBRIDIZATION, NORMALIZATION, PROTEINS, MODULES, VECTORS
UCL classification: UCL
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences > Div of Biosciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences > Div of Biosciences > Genetics, Evolution and Environment
URI: https://discovery.ucl.ac.uk/id/eprint/130779
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