| Budget Amount *help |
¥12,600,000 (Direct Cost: ¥12,600,000)
Fiscal Year 2003: ¥1,800,000 (Direct Cost: ¥1,800,000)
Fiscal Year 2002: ¥1,800,000 (Direct Cost: ¥1,800,000)
Fiscal Year 2001: ¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 2000: ¥5,400,000 (Direct Cost: ¥5,400,000)
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| Research Abstract |
Microarray systems with gene chips(DNA chips) are powerful tools for many researches in post-genome era. However, it is difficult to determine the optimum condition and to get quantitative information by the analysis with a DNA chip because the difference of the thermodynamic stability arisen from one nucleotide difference in a nucleic acid duplex is too small to distinguish the cognate target strand from the others.
The aim of this study is development of "supra-functional" DNA chips on which surface novel functional molecules are attached as an alternative device of the common ones. First of all, thermodynamic stability of triplexes and quadruplexes as well as duplexes was analyzed quantitatively, providing the information for the microarray system that deals with duplex or triplex strands as the analyte. Furthermore, the dependence of the stability was quantified by pH, metal ion, and other co-solutes. For example, a drastic conformational change was found, i.e. antiparallel G-quadruplex structure of d(G_4T_4G_4) changed to parallel orientation in the "cell-mimetic" solution. This phenomenon indicates that the findings in the diluted solution may not be applicable to the DNA chip analysis with the analytes in cell extracts or other concentrated solutions, and therefore, our energy parameters obtained in this study would be the indispensable reference for such a DNA chip system.
Second, the deoxyribozyme developed in my lab was applied to the DNA chip strategy. When the deoxyribozyme chip was used as the sensorchip of the surface plasmon resonance(SPR) analysis, the recognition and the cleavage process of the substrate could be monitored in the sensorgram as expected. This artificial chip is definitely a supra-functional DNA chip exhibits multiple special functions at a time, strict recognition of primary and secondary structure of the substrate, enzymatic cleavage of the substrate, in addition to the normal function of ordinary DNA chips.
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