Detection of (NAD(P)H:Quinone oxidoreductase-1, EC 1.6.99.2) 609C↷T and 465C↷T polymorphisms in formalin-fixed, paraffin-embedded human tumour tissue using PCR-RFLP

  • Authors:
    • Roger M. Phillips
    • Saurajyoti Basu
    • John E. Brown
    • G. Michael Flannigan
    • Paul M. Loadman
    • Sandie W. Martin
    • Brian Naylor
    • Rajiv Puri
    • Tariq Shah
  • View Affiliations

  • Published online on: Thursday, April 1, 2004
  • Pages: 1005-1010
  • DOI: 10.3892/ijo.24.4.1005

Abstract

NQO1 is a cytosolic flavoprotein that plays a dual role in the detoxification of potentially carcinogenic compounds and the bioreductive activation of quinone based anticancer drugs. Two polymorphic variants of NQO1 exist (NQO1*2 and NQO1*3) which cause significant phenotypic reductions in NQO1 protein content and activity. Current methods for detecting NQO1 polymorphisms commonly use PCR-RFLP techniques and have exclusively used DNA isolated from fresh tissues. This study describes a method that is suitable for analysing NQO1 polymorphisms in genomic DNA isolated from formalin-fixed paraffin-embedded tissue. The method utilises two rounds of PCR amplification using a nested primer strategy that generates specific PCR products followed by RFLP analysis using either Hinf1 (for NQO1*2) or Msp1 (for NQO1*3). Whilst existing methods proved unsatisfactory (low product yield and poor specificity), the nested primer strategy produced good quality PCR products suitable for RFLP analysis and genotyping of NQO1*2 and NQO1*3 in archival tissue samples. The ability to utilise the vast archives of human tissue held by pathology laboratories would be of considerable benefit as retrospective studies comparing NQO1 genotype status, patient history and treatment outcomes could be conducted.
Journal Cover

April 2004
Volume 24 Issue 4

Print ISSN: 1019-6439
Online ISSN:1791-2423

2013 Impact Factor: 2.773
Ranked #30/202 Oncology
(total number of cites)

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APA
Phillips, R., Basu, S., Brown, J., Flannigan, G., Loadman, P., Martin, S., Naylor, B., Puri, R., & Shah, T. (2004). Detection of (NAD(P)H:Quinone oxidoreductase-1, EC 1.6.99.2) 609C↷T and 465C↷T polymorphisms in formalin-fixed, paraffin-embedded human tumour tissue using PCR-RFLP. International Journal of Oncology, 24(4), 1005-1010.
MLA
Phillips, Basu, Brown, Flannigan, Loadman, Martin, Naylor, Puri, and Tariq Shah. "Detection of (NAD(P)H:Quinone oxidoreductase-1, EC 1.6.99.2) 609C↷T and 465C↷T polymorphisms in formalin-fixed, paraffin-embedded human tumour tissue using PCR-RFLP." International Journal of Oncology International Journal of Oncology 24.4 (2004): 1005-1010.
Chicago
Phillips, Basu, Brown, Flannigan, Loadman, Martin, Naylor, Puri, and Tariq Shah. "Detection of (NAD(P)H:Quinone oxidoreductase-1, EC 1.6.99.2) 609C↷T and 465C↷T polymorphisms in formalin-fixed, paraffin-embedded human tumour tissue using PCR-RFLP." International Journal of Oncology International Journal of Oncology 24 no. 4 (2004): 1005-1010.