| Project/Area Number |
13470229
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
General surgery
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| Research Institution | HOKKAIDO UNIVERSITY |
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Principal Investigator |
OKADA Futoshi Hokkaido Univ. Institute for Genetic Medicine, Assistant, 遺伝子病制御研究所, 助手 (00250423)
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| Co-Investigator(Kenkyū-buntansha) |
TODO Satoru Hokkaido Univ. Grad. Sch. Of Med., Prof., 大学院・医学研究科, 教授 (60136463)
TADA Mitsuhiro Hokkaido Univ. Institute for Genetic Medicine, Asso. Prof., 遺伝子病制御研究所, 助教授 (10241316)
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| Project Period (FY) |
2001 – 2002
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| Project Status |
Completed (Fiscal Year 2002)
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| Budget Amount *help |
¥11,700,000 (Direct Cost: ¥11,700,000)
Fiscal Year 2002: ¥5,300,000 (Direct Cost: ¥5,300,000)
Fiscal Year 2001: ¥6,400,000 (Direct Cost: ¥6,400,000)
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| Keywords | universal stop codon assay / yeast-based genetic diagnosis / ADE2 / truncating type mutation / tumor suppressor gene / yest-based genetic diagnosis / 腫瘍抑制遺伝子 / 汎用型酵母アッセイ法 / ストップコドンアッセイ |
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| Research Abstract |
In the present study, we aimed to develop a yeast based stop codon assay which is readily applicable to any genes of interest. For this purpose, we studied the following issues : 1) We optimized the method to add sequences to both ends of the PCR-amplified product of tested genes for homologous recombination into the yeast expression vector, and verified efficiency and accuracy of recombination. 2) We found that protein product of some tested gene interfered ADE2 enzymatic activity by hiding the activity center. We added an appropriate spacer sequence to the vector. 3) We verified that the developed stop codon assay could detect truncating type mutations of various genes. Through these process we established and assay called 'universal stop codon assay' (Am J Pathol).
We applied this assay to human cancer sampels, and detected mutation of E-cadherin in ovarian cancers. We studied potential candidate genes for testing by this assay, and identified HOXD3 in lung cancers (Int J Cancer), p73 in brain tumors (Brain Pathol), thymosine-beta4 in soft tissue sarcomas (Am J Pathol), VEGF in osteosarcomas (Br J Cancer), and TGF-beta receptor II in hepatic cancer (Cancer Immunol Immunother). We are now testing these genes with the universal stop codon assay.
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