| Project/Area Number |
10670186
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Human pathology
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| Research Institution | Tokyo Medical University |
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Principal Investigator |
MUKAI Kiyoshi Tokyo Medical University, Department of Pathology, Professor, 医学部, 教授 (20190837)
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| Project Period (FY) |
1998 – 1999
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| Project Status |
Completed (Fiscal Year 1999)
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| Budget Amount *help |
¥2,600,000 (Direct Cost: ¥2,600,000)
Fiscal Year 1999: ¥900,000 (Direct Cost: ¥900,000)
Fiscal Year 1998: ¥1,700,000 (Direct Cost: ¥1,700,000)
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| Keywords | Pathology / PCR / DNA / SSCP / Formalin / Paraffin / ホルマリン固定 / パラフィンブロック / 病理組織標本 |
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| Research Abstract |
It is widely used to extract DNA from formalin-fixed, paraffin-embedded pathology material and use it for genetic analysis by PCR method, However, DNA in paraffin-embedded tissue is modified and degraded during fixation/embedding processes and it is difficult to constantly obtain adequate amount and quality of DNA. Therefore, DNA from fixed tissue cannot be used for Southern blot analysis and the results of PCR analysis are inconsistent. It is the purpose of this study to devise an improved DNA extraction method that can obtain larger amount of good quality DNA from fixed and embedded tissue than the conventional method using phenol/chloroform extraction. Conventional method of phenol/choroloform extraction after xylene deparaffinization and proteinase K digestion was used as a control for the amount and quality of extracted DNA. First, the temperature of deparaffinization or proteinase K digestion was raised. Another method applied microwave irradiation before proteinase K digestion. The last method used heating by microwave irradiation for deparaffinization. The last method consistently extracted more DNA and efficiency of PCR was higher than the other methods. DNA extracted by this method was suitable for PCR-SSCP analysis or direct sequencing of nucleic acids.
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