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Research paper : From basic research on firefly bioluminescence to Product Realization Research (Y. Ohmiya et al.)−234 Synthesiology - English edition Vol.1 No.4 (2009) genes for red and green luciferase of Brazilian railroad worms and found that the 216th amino acid residue in firefly luciferase is involved in determining the color difference of luminescence[3][4]. Railroad worm (or glow-worm) is a luminescent beetle that survives only in Brazil, and expresses a luciferase that produces the strongest red-colored light on earth. Although these research results were new and may help produce biotools for bioluminescence systems, there was no direction that linked the results together. Type 2 Basic Research is not feasible for closing the gap between research and education within the university. In such background, Ohmiya and Nakajima started research at the new National Institute of Advanced Industrial Science and Technology (AIST), where they were given the opportunity to expand the knowledge gained in Type 1 Basic Research to Type 2 Basic Research. Development of biotools using the luminescent color differences started to take shape as an outcome, and a clear goal was set to link bioluminescence to kinetic observations and imaging of biomolecular groups that was being sought in 21st century biosciences.2 What would be developed?In 21st century bioscience after the completion of the Human Genome Project, it is understood that there are limited methods for pursuing a single biomolecule and that innovation would not happen by developing only measurement devices that are extensions of conventional methods. From this background, the “Development of Analysis Technology of Dynamism of Intercellular Network” Project started in 2002, lead by the Ministry of Economy, Trade and Industry, and New Energy and the Industrial Technology Development Organization (NEDO). The objective of this project was to measure efficiently the temporal and special kinetic changes in the intracellular biomolecular network that comprises the foundation of building and functioning of organic tissues in live cells and to establish the technology to allow this functional analysis. The project’s goal was to clarify the information network created by several biomolecules. We participated in this project and suggested the use of luminescent proteins for the development of intracellular labeling technology of multiple types of biomolecules.Our research concept was simple. Whereas traditionally, the amount of light was the only focal point in using firefly luminescence, we looked at the multicolor characteristic of firefly luminescence and studied how to transmit multiple bits of information from the cell. In other words, conventional tools using bioluminescence were an extension of black-and-white television, and we decided to use the color television characteristic, or color difference, to monitor the movement of multiple types of biomolecules in the cell. Our work would connect the results obtained in basic research conducted at the university in the 1990s. Figure 1 is a revised version of one of the slides used in the proposal and describes the research concept and strategy, and the use of luciferase for detecting diverse bioluminescence. Our powerful method was a first-in-the-world result of Type 1 Basic Research showing that the luciferase of railroad worms produces red light from the head and green light from the body.The goal was to develop a multiple drug screening system that would emphasize clearly its difference from conventional technology (Figure 2). The product was ultimately commercialized in April 2006 as “TripLuc” from Toyobo Co., (14)−Fig.1 R&D strategy. Revised version of slide used at NEDO “Intracellular Dynamism Analysis” hearing.The photographs on the left show, from top to bottom, railroad worm, Japanese firefly, sea-firefly, and luminescent jellyfish. The cell function labeling technology that we studied was accomplished using a photomolecular probe as the label that “identifies three or more types of biomolecules without interfering with cell function,” based on luciferases (enzyme or catalyst for luminescence) with different colors obtained from various luminescent organisms.Fig. 2 Example of outcome of R&D. Revised version of slide used at NEDO “Intracellular Dynamism Analysis” hearing.In conventional technology, two gene transcription activities were observed in two steps using two reagents, whereas in the newly developed multireporter assay system, three gene transcription activities could be observed in one step using one reagent. This allows the analysis of several samples at once (high-throughput analysis).Obtain and patent luminescent proteins with different luminescence colorsIdentify three or more types of biomolecules without interfering with cell functionDevelopment of biomolecular labeling technology R&D strategy Railroad worm Sea-fireflyLuminescent jellyfish}Labeled cell function using luciferase genes that produce different colors of luminescence obtained from various luminescent organisms ( )Photo-molecular probe for labeling cell function fireflyImage of expected product realization and current situation “Goal: Development of drug discovery system based on cell function ”Dual reporter assay Development: Multi-reporter assay system Transcriptional activity Reagent 1Reagent Reagent 2Three gene transcription activities with one-step for three reaction using color differences Two gene transcription activities with two-step for two reactions Transcriptional activity

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