| Project/Area Number |
12450375
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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 |
高分子合成
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| Research Institution | RIKEN (2002)
Kyushu University (2000-2001) |
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Principal Investigator |
MAEDA Mizuo RIKEN, Bioengineering Laboratory, Chief Scientist, バイオ工学研究室, 主任研究員 (10165657)
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| Co-Investigator(Kenkyū-buntansha) |
TAKARADA Tohru Graduate School of Engineering, Department of Applied Chemistry, Instructor, 大学院・工学研究院, 助手 (30336010)
村田 正治 九州大学, 大学院・工学研究院, 助手 (30304744)
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| Project Period (FY) |
2000 – 2002
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| Project Status |
Completed (Fiscal Year 2002)
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| Budget Amount *help |
¥12,100,000 (Direct Cost: ¥12,100,000)
Fiscal Year 2002: ¥2,300,000 (Direct Cost: ¥2,300,000)
Fiscal Year 2001: ¥4,100,000 (Direct Cost: ¥4,100,000)
Fiscal Year 2000: ¥5,700,000 (Direct Cost: ¥5,700,000)
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| Keywords | Polymer Micelle / DNA / Poly(N-isopropylacrylamide) / Gene Diagnosis / Single Nucleotide Mutation / SNP / Nanoparticle / Oncogene / ポリ(N-イソプロピルアクリルアミド) / 塩析 / コロイド分散液 |
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| Research Abstract |
A single nucleotide mutation on certain genes can cause heritable disorders and cancers. Consequently the development of a simple and rapid detection method for single nucleotide difference has been one of the most important subjects in analytical biochemistry. In this study, we applied a salting out-induced turbidity change of polymer micelle dispersion to single nucleotide difference assay.
The polymer micelle comprises a hydrophobic core of poly(N'-isopropylacrylamide) (polyNIPAAm) and a hydrophilic shell of oligodeoxyribonucleotide (ODN : 9 mer). The micelle was constructed by self-assembly of amphiphilic copolymers of ODN derivative and NIPAAm. The amphiphilic copolymer was synthesized through copolymerization between NIPAAm and ODN macromonomer, which was obtained by coupling reaction between methacryloyloxysuccinimide and ODN having amino-hexyl linker at its 5'-end. When the copolymer solution was incubated above the phase transition temperature of polyNIPAAm, the polymer micelle was spontaneously formed and kept dispersed.
The polymer micelles aggregated rapidly when the complementary ODN (9 mer) was added into the dispersion. In contrast, they kept dispersed in the presence of the point-mutated ODN (9 mer). Moreover, we have applied an extended method for the duplex formation to detect single nucleotide mutation of longer sample ODN (39 mer). In this method, the probe ODN (10 mer) and the auxiliary ODN (20 mer) were added to complement the surplus part (30 bases) of the sample ODN. The detection part of the sample ODN is 10 bases wide, which is complementary to the probe ODN. Using this method, we can detect the single nucleotide mutation of the longer sample ODN. These distinct phenomena may be applied for a DNA discrimination system in gene diagnosis.
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