2014 Fiscal Year Final Research Report
Analysis of DNA metabolic reactions based on single molecule visualization method.
| Project/Area Number |
24770164
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| Research Category |
Grant-in-Aid for Young Scientists (B)
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| Allocation Type | Multi-year Fund |
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| Research Field |
Molecular biology
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| Research Institution | Gunma University |
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Principal Investigator |
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| Project Period (FY) |
2012-04-01 – 2015-03-31
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| Keywords | １分子観察 / 計測技術 / 可視化技術 / ＤＮＡ代謝反応 / ＤＮＡ複製 / ＤＮＡポリメラーゼ / ヌクレアーゼ / ＤＮＡ結合タンパク質 |
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| Outline of Final Research Achievements |
DNA synthesis and DNA digestion reactions were studied using direct single-molecule observations in microflow channels. In the channels, ssλDNA shape was controlled as relaxed and stretched form by adjusting buffer flow rate, and DNA synthesis reactions by Klenow Fragment (3’-5’ exo-) were observed. Analysis of the synthetic reaction rates demonstrated that the DNA synthesis reaction rate of stretched ssλDNA was approximately 75% higher than that of random coiled ssλDNA . Also, DNA digestion reactions by T7 Exonuclease (T7 Exo) were directly observed both under pulse-chase conditions and under continuous buffer flow conditions with T7 Exo. Under pulse-chase conditions, the double-stranded regions of λDNA monotonously shortened by a DNA digestion reaction with a single T7 Exo molecule. Under continuous buffer flow conditions with T7 Exo, some pauses were observed during a DNA digestion reaction and double-stranded regions shortened linearly except during these pauses.
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| Free Research Field |
分子生物学
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