2001 Fiscal Year Final Research Report Summary
Real-time monitoring of gene expression by bioluminescence
| Project/Area Number |
11558089
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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 |
Cell biology
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| Research Institution | Nagoya University |
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Principal Investigator |
KONDO Takao Nagoya University Graduate School of Science, Professor, 大学院・理学研究科, 教授 (10124223)
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| Co-Investigator(Kenkyū-buntansha) |
ISHIURA Masahiro Nagoya University Gene Research Center, Professor, 遺伝子実験施設, 教授 (20132730)
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| Project Period (FY) |
1999 – 2001
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| Keywords | cyanobacteria / bioluminescence / real-time monitoring system / rail-road worms / luciferase / gene expression |
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| Research Abstract |
Analysis of gene expression is essential for all aspects of biology today. Real-time monitoring of gene expression from living cells is very powerful tool to dissect molecular mechanisms for many biological phenomena. To apply this technique to wide range of cells, a stable and versatile method need to be developed. In this project, we analyzed many technical problem that the in vivo real-time method should address.
Bioluminescence from luciferase protein in the cell is affected by various factors, including subtrate, stability of enzyme, energy supply, etc. To avoid these factors, we applied two luciferase system of railroad worm to this purpose. This insect have two similar luciferase genes. While two luciferases share similar characteristics, waveform of bioluminescence is quite different each other (540 and 630 nm). With suppy of thses genes, we introduce them to cyanobacteria under different promoters. Then we developed automated bioluminescence detection system based on two photomultiplier tube and interference filters of different wave length. We finally succeeded to obtain cyanobacteria that contains two luciferase reporter and confirmed that siginals from two promoters can be detected without significant cross-talk. Therefore, now we can monitor expressions of two gene concurrently. With this method, we will enhance reliability to bioluminescence signals and evaluate gene expression in higher precision. Application of these reproter system can be expanded to various biological system to obtain reliable monitor of gene expression.
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[Publications] Iwasaki, H., Williams, S.B., Kitayama Y., Ishiura, M., Golden, S.S. and T. Kondo: "A KaiC-interacting sensory histidine kinase, SasA, necessary to sustain robust circadian oscillation in cyanobacteria."Cell. 101. 223-233 (2000)
Description
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