September 1997, Volume 3, Issue 3, Pages 157 – 161

Paper

Paraffin-embedded tissue as a source of RNA for gene expression analysis in oral malignancy MT Cairns1, S Church1, PG Johnston1, KV Phenix2 & JJ Marley2 1Department of Oncology, The Queen’s University of Belfast, Belfast City Hospital Tower, Lisburn Rd, Belfast BT9 7AB, UK     2School of Clinical Dentistry, The Queen’s University of Belfast, Royal Group of Hospitals, Grosvenor Rd, Belfast BT12 6BP, UK

Keywords

RNA;   RT-PCR;   oral squamous cell carcinoma

Abstract

OBJECTIVE: To assess the feasibility of using archival oral mucosal tissue to examine gene expression at the ribonucleic acid (RNA) level. MATERIALS AND METHODS: We describe the isolation of RNA from 8 µm sections of formalin-fixed paraffin-embedded oral mucosal tissue. RNA was reverse transcribed and three candidate genes amplified by polymerase chain reaction (PCR). The ribosomal protein S14 gene is a housekeeping gene which has been used as an internal standard in several quantitative PCR protocols. The thymidine kinase (TK) gene is expressed at low levels in most tissues and, with a well-documented genomic organisation, is a useful tool for discrimination between genomic DNA and cDNA. The RI gene is reported to be overexpressed in many cancer cell lines, in malignant tissue and in vitro transformed cells. RESULTS: The S14 gene, the TK gene and the RI gene of the cAMP-dependent protein kinase (PKA) were amplified successfully from formalin-fixed paraffin-embedded tissue sections. The TK primer pair is a useful additional tool in the unambiguous identification of RNA-derived species. CONCLUSION: RNA suitable for reverse transcribed (RT)-PCR was extracted from archival oral mucosal tissue. This should permit rapid sequence analysis of transcribed tumor suppressor genes and oncogenes in this material. Furthermore, the RT-PCR approach described may allow quantification of gene expression in oral mucosal archival material processed in a standard fashion.

Received 4 September 1996; Revised 22 May 1997; Accepted 14 July 1997

^© Macmillan Publishers Ltd 1997