2000 Fiscal Year Final Research Report Summary
Davelopment of Method to Analyse the Function of Regulatory Proteins in Single by Cells by Gene Handling Technique.
| Project/Area Number |
11559016
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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 | 展開研究 |
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| Research Field |
広領域
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| Research Institution | Waseda University |
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Principal Investigator |
ISHIWATA Sinichi Waseda University, School of Scienc, and Engineering, Professor, 理工学部, 教授 (10130866)
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| Co-Investigator(Kenkyū-buntansha) |
FUJITA Hideaki Japan Society for the Promotion of Science Research Fellow, 特別研究員 (50318804)
OKANO Kazunori Hitachi, Ltd, Central Research Laboratory, Senior Research Scientist, 中央研究所, 主任研究員
YASUDA Kenji The Uhiversity of Tokyo, Graduate School of Arts and Sciences, Associate Professor, 大学院・総合文化研究科, 助教授 (20313158)
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| Project Period (FY) |
1999 – 2000
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| Keywords | Caged DNA / Teaperature-pulse unicroscopy / DNA tip / POR method / Myoblast cell / GFP-actin / Actin filament / Stress sensor |
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| Research Abstract |
Recovery of DNA fragment from a DNA probe array : We have developed a DNA preparation method using a DNA probe array that utilizes photo-thermal denaturation to recover specific DNA. The protocol for preparing DNA probe array was investigated to realize the stable immobilization of DNA probes on the surface of solid support. We used a glass plate coated with Cr (5-10 nm thick). The Cr surface was modified with 3-glysidoxypropyltrimethoxysilane to introduce active residues that can couple with amino residues at the 5' terminal of DNA probes. The Cr surface acts as a photo-thermal transducer. The preparation method of DNA uses infrared (1053-nm) laser irradiation to thermally denature and release DNA immobilized in a specific area of a DNA probe array. Different DNA fragments fixed in place on the DNA probe array could be separately recovered. There were enough quantities of recovered DNA that can be amplified by using PCR. Trial to synthesize caged DNA : Aiming at synthesizing caged DNA, we tried to chemically modify an amino-residue on DNA with 4, 5-diraethoxy-2-nitrobenzyl bromide (DMNBB). Although the synthesis of chemically modified DNA once appeared to be confirmed by using PCR method, we finally reached a conclusion that the amino-residue on DNA is not reactive to the chemical modification. Expression of GFP-actin in cultured embryonic heart cells : We could confirm that GFP-actin was expressed in cultured embryonic heart cells under fluorescence microscopy. Although we may have not be able to achieve the original goal, the results obtained in this project must be, we believe, very useful for planning a future project.
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