Improvement in a E. coli host cell for DNA cloning of long DNA fragments.
| Project/Area Number |
18K19193
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| Research Category |
Grant-in-Aid for Challenging Research (Exploratory)
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| Allocation Type | Multi-year Fund |
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| Review Section |
Medium-sized Section 38:Agricultural chemistry and related fields
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| Research Institution | National Institute of Genetics |
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Principal Investigator |
NIKI Hironori 国立遺伝学研究所, 遺伝形質研究系, 教授 (70208122)
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| Project Period (FY) |
2018-06-29 – 2021-03-31
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| Project Status |
Completed (Fiscal Year 2020)
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| Budget Amount *help |
¥6,240,000 (Direct Cost: ¥4,800,000、Indirect Cost: ¥1,440,000)
Fiscal Year 2019: ¥3,640,000 (Direct Cost: ¥2,800,000、Indirect Cost: ¥840,000)
Fiscal Year 2018: ¥2,600,000 (Direct Cost: ¥2,000,000、Indirect Cost: ¥600,000)
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| Keywords | 形質転換体 / クローニング / オメガ因子 / RNAポリメラーゼ / iVEC / ゲノム / 遺伝子組換え / 形質転換 / DNA クローニング / 大腸菌宿主 / ゲノム合成 |
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| Outline of Final Research Achievements |
In the in vivo cloning (iVEC) method, a recombinant clone can be obtained simply by transforming the vector DNA and the cloned DNA, which are PCR products with homologous sequences, into an Escherichia coli cell. This technique has the advantage that seamless cloning can be performed inexpensively and quickly. We conducted a genetic elucidation of the molecular mechanism of the in vivo cloning and aimed to develop an Escherichia coli strain with high transformation efficiency to improve this activity. In the search for genes related to high transformation efficiency, genes involved in transformation were found. We found the rpoZ gene as a candidate gene that improves transformation efficiency. The rpoZ gene regulates gene groups whose expression is involved in transformation efficiency.
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| Academic Significance and Societal Importance of the Research Achievements |
in vivoクローニング(iVEC)法の分子メカニズムの遺伝学的な解明を行った。in vivoクローニングの分子メカニズムには3’-5’エンドヌクレアーゼXthAが必須であった。XthAはDNAの3’末端から二本鎖DNAを一本鎖DNAが露出し、相同な一本鎖 DNA同士がアニーリングをさせる活性であることを明らかにした。これを元により高効率な方法への改善が期待され、形質転換の向上を同時に進めることでさらなる組換えDNA技術の発展が見込まれる。
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Report
(4 results)
Research Products
(3 results)
