人工合成細胞工廠研究

張衛(wèi)文 陳磊 周杰 馬鋼 曹旭鵬

摘 要：成功開發(fā)了高效、穩(wěn)定的藍細菌遺傳改造工具。使用該工具可以實現(xiàn)對不同基因進行高表達或低表達；通過理性設計基因組插入位點，實現(xiàn)對藍細菌內容碳流進行重新分配。該工具正在藍細菌創(chuàng)建生物合成途徑中測試。成功創(chuàng)建了從CO2到異丙醇的生物合成途徑，實現(xiàn)利用CO2生物合成異丙醇。突變株生理特性測試及優(yōu)化正在進行中。構建了針對雙組件相應調控蛋白，轉錄調控因子，以及細胞小RNA等的藍細菌刪除株共273株，以及18株含有不同功能基因的藍細菌過表達株。使用基于96孔板的底盤檢測技術，對構建獲得的藍細菌底盤進行了生物產品耐受性（針對乙醇、丁醇、3羥基丙酸3HP等）以及環(huán)境脅迫因子耐受（氮源脅迫、金屬離子脅迫、高鹽脅迫等）進行了系統(tǒng)的分析，獲得多株對于各種脅迫因素以及生物產品發(fā)生抗性改變的底盤細胞。確定了一個與集胞藻脂肪酸代謝相關的轉錄因子，其在脂肪酸代謝網絡中的位置及功能的確定還在進一步分析中。完成了兩個重要蛋白的晶體結構解析。

關鍵詞：人工合成細胞工廠 光合藍細菌 底盤細胞

Abstract：A highly efficient and stable genetic transformation tool for cyanobacteria was successfully developped of. High or low expression of different genes can be achieved by use of this tool. A biosynthetic pathway from CO2 to isopropyl alcohol was successfully created. 273 cyanobacteria gene knockout mutants were constructed for the corresponding two- component regulatory proteins， transcription factors， or small RNAs. 18 cyanobacteria overexpression strains containing different functional genes were also constructed. Phenotype analysis was performed and a series of mutants that are tolerant to ethanol， butanol ， 3HP and some environmental stress factors such as nitrogen stress， metal ion stress， high salt stress， were identified. A fatty acid metabolism related transcription factor in Synechocystis was identified. Crystal structure analysis of two important proteins was completed.

Key Words：Synthetic cell factories；Photosynthetic cyanobacteria；Chassis cell

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