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Comparison of the DNA Extraction Methods for Polymerase Chain Reaction Amplification from Formalin-Fixed and Paraffin-Embedded Tissues

Sato, Yuichi Ph.D.; Sugie, Rikako M.S.; Tsuchiya, Benio Ph.D.; Kameya, Toru M.D.; Natori, Michiya M.D.; Mukai, Kiyoshi M.D.

Diagnostic Molecular Pathology: December 2001 - Volume 10 - Issue 4 - pp 265-271
Original Articles
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To obtain an adequate quality and quantity of DNA from formalin-fixed and paraffin-embedded tissue, six different DNA extraction methods were compared. Four methods used deparaffinization by xylene followed by proteinase K digestion and phenol-chloroform extraction. The temperature of the different steps was changed to obtain higher yields and improved quality of extracted DNA. The remaining two methods used microwave heating for deparaffinization. The best DNA extraction method consisted of deparaffinization by microwave irradiation, protein digestion with proteinase K at 48°C overnight, and no further purification steps. By this method, the highest DNA yield was obtained and the amplification of a 989-base pair β-globin gene fragment was achieved. Furthermore, DNA extracted by means of this procedure from five gastric carcinomas was successfully used for single strand conformation polymorphism and direct sequencing assays of the β-catenin gene. Because the microwave-based DNA extraction method presented here is simple, has a lower contamination risk, and results in a higher yield of DNA compared with the ordinary organic chemical reagent–based extraction method, it is considered applicable to various clinical and basic fields.

From the Department of Molecular Diagnostics, School of Allied Health Sciences (Y.S., R.S., B.T.) and Department of Pathology, School of Medicine (T.K.), Kitasato University, Kanagawa; Department of Clinical Research (M.N.), National Okura Hospital, Tokyo; and Department of Pathology (K.M.), Tokyo Medical University, Tokyo, Japan.

Address correspondence and reprint requests to Dr. Yuichi Sato, Department of Molecular Diagnostics, School of Allied Health Sciences, Kitasato University, 1–15–1 Kitasato, Sagamihara-shi, Kanagawa 228–8555, Japan; email: yuichi@med.kitasato-u.ac.jp.

This work was supported in part by a grants-in-aid for Pediatric Research (10C-2) and for Cancer Research (10–28) from the Ministry of Health and Welfare of Japan.

© 2001 Lippincott Williams & Wilkins, Inc.