1991 Fiscal Year Final Research Report Summary
Study on Meta-stable Secondary Structures of Nucleic Acids in Solution
| Project/Area Number |
01480543
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| Research Category |
Grant-in-Aid for General Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
生物物性学
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| Research Institution | Saitama University |
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Principal Investigator |
HUSIMI Yuzuru Saitama Univ., Dept. of Environ. Chem., Prof., 工学部, 教授 (80011641)
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| Co-Investigator(Kenkyū-buntansha) |
SUZUKI Miho Saitama Univ., Dept. of Environ. Chem., Res. Assoc., 工学部, 助手 (60222064)
MIYASHITA Akira Saitama Univ., Dept. of Appl. Chem., Assoc. Prof., 工学部, 助教授 (90132729)
NISHIGAKI Koichi Saitama Univ., Dept. of Environ. Chem., Assoc. Prof. (Head investigator in 1989-, 工学部, 助教授 (10107378)
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| Project Period (FY) |
1989 – 1991
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| Keywords | Nucleic acid / Secondary structure / Meta-stable secondary structure / Denaturing gel electrophoresis / TGGE / Random PCR / Primer extension reaction / DNA profiling |
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| Research Abstract |
The knowledge of the solution structure of nucleic acids is the most important for the understanding of their function. Hitherto, the main approaches to this problem were limited to the analyses of the stable structures, which is quite natural because the stable ones could be more easily attacked by the conventional methods such as X-ray cryitallography. This study developed the analysis of metastable structures of nucleic acids, which are believed to take significant part in the ir functions. The following results were obtained : 1) Development of precise denaturing gradient gel electrophoresis, application to obtaining information on solution structures of nucleic acids and advancing of the developed techniques : Denaturing gradient gel electrophoresis was sophisticated into the highly reproducible, objective and informative system, which gave the way to grasp the structural polymorphism and their dynamics as a function of denaturing effect. 'Standardized mobility profile', which is
… More
specific to a molecule, is an important property, by which we can not only separate and identify molecules but also obtain the basic data of structure-mobility relationship. The experiment on ds-DNAs using precise TGGE clarified that structure-mobility relationship of them might be obtainable. Experimental data on strand separation and cooperative melting became also obtainable by this method. This technique was developed to 'DNA profiling', which was a new technique combined with random PCR and was shown to analyse directly a huge genomic DNA. 2) The other technical developments and the phenomenological findings of solution structures of nucleic acids : We developed the 'priming activity assay method' for the analysis of the solution structure of ss-nucleic acids, which was also exploited into a micro ISFET pH-sensor system. This method enabled us to measure the amount of meta-stable intra- and intermolecular secondary structures while we had preliminarily succeeded in predicting those structures by way of computer analysis. In regard to these, we could also establish 'primer-stopper method', 'peculiar cleavage of ssDNA by a cooperative action of a restriction enzyme and an oligonucleotide' and the others. Less
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[Publications] NISHIGAKI, K., MIURA, T., TSUBOTA, M., SUTOH, A., AMANO, N. and HUSIMI, Y.: "Structual Analysis of Nucleic Acids by Precise Denaturing Gradient Gel Electrophoresis : II. Application to the Analysis of Subtle and Drastic Mobility Changes of Oligo- and Polynucleotide" J. Biochem. 111. 151-156 (1992)
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