| Co-Investigator(Kenkyū-buntansha) |
HAGIYA Masami The University of Tokyo, Graduate School of Information Sciences and Technology, Professor, 大学院・情報理工学系研究科, 教授 (30156252)
SUYAMA Akira The University of Tokyo, Graduate School of Arts and Sciences, Professor, 大学院・総合文化研究科, 教授 (90163063)
KOMIYA Ken The University of Tokyo, Tokyo Institute of Technology Interdisciplinary Graduate School of Science and Engineering, Assistant Professor, 大学院・総合理工学研究科, 助手 (20396790)
坂本 健作 東京大学, 大学院・理学系研究科, 助手 (50240685)
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| Budget Amount *help |
¥16,500,000 (Direct Cost: ¥16,500,000)
Fiscal Year 2005: ¥1,700,000 (Direct Cost: ¥1,700,000)
Fiscal Year 2004: ¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 2003: ¥11,200,000 (Direct Cost: ¥11,200,000)
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| Research Abstract |
In Whiplash PCR (WPCR), autonomous molecular computation is implemented in vitro by the recursive, self-directed polymerase extension of a DNA hairpin mixture. In principle, WPCR is capable of solving instances of various NP-complete problems, and of supporting both in vitro evolutionary computation (EWPCR), and programmable protein evolution (XWPCR), via combination with RNA-protein fusion. Unfortunately, a practical barrier for scaling is a systematic tendency for strands to self-poison via back-hybridization (BH), which cannot be eliminated by either reaction condition optimization or strand encoding.
In this project, the impact of BH on WPCR and variants has been characterized experimentally, and the underlying computational process has been re-designed and optimized to support in vitro protein evolution via XWPCR. Major achievements include : (1)experimental validation of our thermodynamic model of process efficiency ; (2)development of a coupled equilibrium model for high-fidelity
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Tag-Antitag system design to assist strand anchorage ; (3)implementation of the software tool, NucleicPark, to evolve DNA words for high-fidelity computation and anchorage ; and, (4)development and experimental validation of Displacement WPCR (DWPCR), a substantially improved architecture.
A critical advance for WPCR, DWPCR employs targeted strand displacement and site protection by DNA polymerase, rather than PNA-targeting, to eliminate BH, achieving an efficiency approaching unity. DWPCR also eliminates thermal cycling, allowing in vivo applications. In contrast with PWPCR, target sequences for primer extension are segregated from the computation, so that the destabilization of shuffled proteins predicted for PWPCR-based XWPCR, due to translation of targeting artifacts is absent. The accompanying architectural freedom also permits design for XWPCR-based exon shuffling (i.e., protein module rather than pseudo-module shuffling), allowing XWPCR to more closely imitate natural evolving systems. DWPCR and the developed models and tools are expected to provide an experimentally validated platform for efficient molecular computation, specifically optimized for evolutionary programming via DWPCR-based EWPCR and XWPCR Less
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