至今在藍綠菌細胞中為了能夠系統化得設計基因電路，建立啟動子-核糖體的元件庫，以因應各種廣泛的應用仍是一件吸引人的事。在藍綠菌細胞 PCC 7942 中，為了能夠系統化得設計基因電路，我們建立了三種啟動子-核糖體結合位元件庫，即連續表現型、抑制子調控型及促進子調控型。將工程化基因電路轉殖到藍綠菌中，分別提供了這三種啟動子-核糖體結合位庫的建立流程圖，根據實驗中量測到啟動子-核糖體的強度和動態模型，啟動子-核糖體被視為一種特性組件，再來透過系統鑑別參數的技術將生物元件特性化，並且調控不同種類的生物組件，用於基因電路設計以符合在藍綠菌細胞中的期望行為。在藍綠菌細胞中，展現了設計的示範例子關於細胞族控制，說明了設計流程根據所提出的啟動子-核糖體搜尋方法，並且結合活體細胞做驗證。為了使設計更加方便，利用系統化方法來控制從元件庫中選擇出的核糖體元件強度，以滿足在藍綠菌細胞中的行為。因此我們相信所提出系統化的工程設計方法，能提供快速發展的合成生物領域，讓使用者能選擇最佳的元件以實現各種的應用。

At present, it is still appealing to have promoter-RBS libraries for systematic design of synthetic genetic circuit in Cyanobacteria for a variety of application. In the Cyanobacterium Synechococcus sp. strain PCC 7942, we set up three kinds of promoter-RBS libraries, i.e., constitutive, repressor-regulated and activator-regulated promoter-RBS libraries, for systemic design of synthetic gene circuits in Cyanobacteria. In order to engineer synthetic gene circuit into Cyanobacterium Synechococcus sp. PCC 7942, we provide the flow chart to constructing constitutive, repressor-regulated, and activator-regulated promoter-RBS libraries based on promoter-RBS intensities through experimental data, i.e., , , and , respectively. By the parameter identification technique based on dynamic model, a promoters-RBS is regarded as a characteristic part to be selected for gene circuit design with a desired behavior in Cyanobacteria, and regulated by different biological parts in blue-green algae. A design example with desired cell population control is given to illustrate the design procedure of synthetic circuit in Cyanobacteria by the proposed promoter-RBS library-search method and is validated by experimental data in vivo. For the convenience of design, we provide a systematic method for a genetic circuit to control part’s intensity to achieve some desired behavior in Cyanobacterium Synechococcus sp. PCC 7942. Consequently, we believe that the proposed library-based search method can help user select an adequate set of promoter-RBS parts to realize various kinds of application in rapidly developed synthetic biology.

摘要 i
Abstract ii
誌謝 iii
Content iv
List of Figures vii
List of Tables viii
Introduction 1
The flow chart of building Promoter-RBS Libraries for synthetic gene circuit in Cyanobacteria 5
2.1. Characteristic Indexes of Promoter-RBS libraries based on promoter-RBS intensity 5
2.2. Genetic engineering of Cyanobacteria 7
2.3. Dynamic model for protein expression of synthetic gene circuit in Cyanobacteria 9
2.4.The flow chart of building constitutive promoter-RBS libraries in Cyanobacteria
11
2.5. The flow chart of building repressor-regulated promoter-RBS libraries 13
2.6. The flow chart of building activator-regulated promoter-RBS libraries 18
Design of function gene circuit for cell population control in Synechococcus sp. PCC 7942 23
3.1. The introduction of Bacteriorhodopsin (BR) and cyanobacterial ictB gene 23
3.2. Dynamic model of control genetic circuit for cell population control of Cyanobacteria 25
3.3. Design specifications and procedure of population control genetic circuit for cell population control in Cyanobacteria 27
3.4. Validation of cell population control genetic circuit in Cyanobacteria 30

Discussion 32
Conclusion 35
Reference 36
Figures 41
Tables 56
Material and Method 61

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