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Whole-genome microarrays of fission yeast: characteristics, accuracy, reproducibility, and processing of array data
- Source :
- BMC Genomics, BMC Genomics, Vol 4, Iss 1, p 27 (2003)
- Publication Year :
- 2003
- Publisher :
- BioMed Central, 2003.
-
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.<br />RIGHTS : This article is licensed under the BioMed Central licence at http://www.biomedcentral.com/about/license which is similar to the 'Creative Commons Attribution Licence'. In brief you may : copy, distribute, and display the work; make derivative works; or make commercial use of the work - under the following conditions: the original author must be given credit; for any reuse or distribution, it must be made clear to others what the license terms of this work are.
- Subjects :
- Normalization (statistics)
Microarray
lcsh:QH426-470
PROTEINS
lcsh:Biotechnology
Genes, Fungal
Computational biology
Biology
Genome
03 medical and health sciences
vectors
SCHIZOSACCHAROMYCES-POMBE
lcsh:TP248.13-248.65
Schizosaccharomyces
Genetics
hybridization
030304 developmental biology
GENE-EXPRESSION
DNA Primers
Fluorescent Dyes
Oligonucleotide Array Sequence Analysis
0303 health sciences
Gene Expression Profiling
030302 biochemistry & molecular biology
DNA microarray
Reproducibility of Results
Genomics
biology.organism_classification
MEIOSIS
Gene expression profiling
lcsh:Genetics
normalization
Coding strand
Schizosaccharomyces pombe
MODULES
Biotechnology
Research Article
Subjects
Details
- Language :
- English
- ISSN :
- 14712164
- Volume :
- 4
- Database :
- OpenAIRE
- Journal :
- BMC Genomics
- Accession number :
- edsair.doi.dedup.....fc7dfd9937dfb15337d4289ed35aa759