| Summary: | 碩士 === 國立交通大學 === 生物科技學系 === 100 === Genetic engineering with recombinant DNA is a powerful and widespread technology that enables researchers to redesign life forms by modifying DNA fragments. Programming and controlling cell behavior requires fine control the protein expression levels. Previous studies provide several methods to predict the transcription rates of promoters and translation rates of ribosome binding sites (RBSs) respectively. However, the protein expression level with time is hard to predict properly by those methods. To overcome this problem, we selected four promoters and three RBSs with different regulation strength and constructed 12 protein expression devices which combine promoter, RBS and green fluorescent protein (GFP) in Escherichia coli. The GFP expression levels with time were measured using a flow cytometry, and the experimental data can used to characterize a protein expression rate of a protein expression device which contains a promoters and a RBS. A dynamic model that captured the experimentally observed differences for each protein expression device was developed in this study. Using this model, we can define the protein expression rate in the different E. coli population density for the protein expression device. To demonstrate reverse engineering, this model was used to predict the protein expression level in repressor-controlled genetic circuits, and the experimental results consistent with our prediction. Thus, this model enable us to rational connect a promoter and a RBS to obtain a target protein expression level in a complex genetic circuits. Our method can quantitative the protein expression rate at different E.coli population density and can implement a genetic circuit with desired function in E.coli.
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