Microarray validation: factors influencing correlation between oligonucleotide microarrays and real-time PCR

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• 作者：Jeanine S. Morey (1)
  James C. Ryan (1)
  Frances M. Van Dolah (1)
  
• 关键词：Polymerase Chain Reaction ; Microarray Analysis ; Gene Expression ; Nucleic Acid Amplification Techniques ; Reverse Transcriptase Polymerase Chain Reaction ; RNA
• 刊名：Biological Procedures Online
• 出版年：2006
• 出版时间：December 2006
• 年：2006
• 卷：8
• 期：1
• 页码：175-193
• 全文大小：1141KB
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• 作者单位：Jeanine S. Morey (1)
  James C. Ryan (1)
  Frances M. Van Dolah (1)
  
  1. NOAA Marine Biotoxins Program, Center for Coastal Environmental Health and Biomolecular Research, 219 Fort Johnson Rd., 29412, Charleston, SC, USA
  
• ISSN：1480-9222

文摘

Quantitative real-time PCR (qPCR) is a commonly used validation tool for confirming gene expression results obtained from microarray analysis; however, microarray and qPCR data often result in disagreement. The current study assesses factors contributing to the correlation between these methods in five separate experiments employing two-color 60-mer oligonucleotide microarrays and qPCR using SYBR green. Overall, significant correlation was observed between microarray and qPCR results (ρ=0.708, p<0.0001, n=277) using these platforms. The contribution of factors including up -vs. down-regulation, spot intensity, ρ-value, fold-change, cycle threshold (Ctsub>), array averaging, tissue type, and tissue preparation was assessed. Filtering of microarray data for measures of quality (fold-change and ρ-value) proves to be the most critical factor, with significant correlations of ρτ;0.80 consistently observed when quality scores are applied.