Assessment of the sensitivity and specificity of oligonucleotide (50mer) microarrays

To examine the utility and performance of 50mer oligonucleotide (oligonucleotide probe) microarrays, gene-specific oligonucleotide probes were spotted along with PCR probes onto glass microarrays and the performance of each probe type was evaluated. The specificity of oligonucleotide probes was stud...

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Bibliographic Details
Published in:Nucleic acids research Vol. 28; no. 22; pp. 4552 - 4557
Main Authors: Kane, Michael D., Jatkoe, Timothy A., Stumpf, Craig R., Lu, Jia, Thomas, Jeffrey D., Madore, Steven J.
Format: Journal Article
Language:English
Published: England Oxford University Press 15.11.2000
Subjects:
Animals
Bacillus subtilis - genetics
Base Sequence
DNA Probes
Gene Expression Regulation, Bacterial
Genes, Bacterial - genetics
microarrays
Molecular Sequence Data
Oligonucleotide Array Sequence Analysis - methods
Oligonucleotides - genetics
Rats
RNA, Messenger - genetics
RNA, Messenger - metabolism
Sensitivity and Specificity
ISSN:0305-1048, 1362-4962, 1362-4962
Online Access:Get full text
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Summary:To examine the utility and performance of 50mer oligonucleotide (oligonucleotide probe) microarrays, gene-specific oligonucleotide probes were spotted along with PCR probes onto glass microarrays and the performance of each probe type was evaluated. The specificity of oligonucleotide probes was studied using target RNAs that shared various degrees of sequence similarity. Sensitivity was defined as the ability to detect a 3-fold change in mRNA. No significant difference in sensitivity between oligonucleotide probes and PCR probes was observed and both had a minimum reproducible detection limit of ∼10 mRNA copies/cell. Specificity studies showed that for a given oligonucleotide probe any ‘non-target’ transcripts (cDNAs) >75% similar over the 50 base target may show cross-hybridization. Thus non-target sequences which have >75–80% sequence similarity with target sequences (within the oligonucleotide probe 50 base target region) will contribute to the overall signal intensity. In addition, if the 50 base target region is marginally similar, it must not include a stretch of complementary sequence >15 contiguous bases. Therefore, knowledge about the target sequence, as well as its similarity to other mRNAs in the target tissue or RNA sample, is required to design successful oligonucleotide probes for quality microarray results. Together these results validate the utility of oligonucleotide probe (50mer) glass microarrays.
Bibliography:Received July 5, 2000; Revised and Accepted September 21, 2000.
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To whom correspondence should be addressed at present address: Pfizer Global Research and Development, 2800 Plymouth Road, Ann Arbor, MI 48105, USA. Tel: +1 734 622 1782; Fax: +1 734 622 5970; Email: steven.madore@pfizer.com Present address: Michael D. Kane, Genomic Solutions Inc., 4355 Varsity Drive East, Ann Arbor, MI 48108, USA
ISSN:0305-1048
1362-4962
1362-4962
DOI:10.1093/nar/28.22.4552