Dextran sodium sulfate inhibition of real-time polymerase chain reaction amplification: A poly-A purification solution

T. A. Kerr, M. A. Ciorba, H. Matsumoto, V. R.T. Davis, J. Luo, S. Kennedy, Y. Xie, A. Shaker, B. K. Dieckgraefe, N. O. Davidson

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23 Scopus citations

Abstract

Background: Dextran sulfate sodium (DSS) induces experimental colitis and promotes colitis-associated cancer in rodents. Here we document potent inhibition of real-time quantitative polymerase chain reaction (qPCR) using cDNA from DSS-exposed mouse tissues, which complicates gene expression analysis. Methods: We characterize DSS inhibition of qPCR in-vitro and in a wide array of murine tissues following ingestion of DSS. We examine different approaches to RNA purification prior to cDNA synthesis in order to optimize real-time polymerase chain reaction amplification and gene expression analysis. Results: DSS inhibits qPCR amplification of cDNA between 1 and 10 nM. Orally administered DSS interferes with qPCR amplification of cDNA derived from multiple tissues. Poly-A purification of DSS-exposed RNA allows reliable and cost-effective gene expression analysis in DSS-exposed tissue. Conclusions: DSS is a potent inhibitor of real-time qPCR amplification and interferes with tissue-specific gene expression analysis in DSS-exposed mice. Poly-A purification of tissue-derived RNA results in reliable and cost-effective gene expression analysis in DSS-exposed mice.

Original languageEnglish (US)
Pages (from-to)344-348
Number of pages5
JournalInflammatory Bowel Diseases
Volume18
Issue number2
DOIs
Publication statusPublished - Feb 1 2012

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Keywords

  • colitis
  • colitis-associated cancer
  • dextran sodium sulfate
  • qPCR

ASJC Scopus subject areas

  • Immunology and Allergy
  • Gastroenterology

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