A method for Absolute Protein Expression Quantity Measurement Employing Insulator RiboJ

Hongbin Yu; Zheng Wang; Hanyue Xu; Jiusi Guo; Qingge Ma; Xiangxu Mu; Yunzi Luo

doi:10.1016/j.eng.2018.09.012

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Engineering ›› 2018, Vol. 4 ›› Issue (6) : 881-887. DOI: 10.1016/j.eng.2018.09.012

Research

Research Synthetic Biology—Article

A method for Absolute Protein Expression Quantity Measurement Employing Insulator RiboJ

Hongbin Yu^a ,
Zheng Wang^a^,^c ,
Hanyue Xu^a ,
Jiusi Guo^a^,^c ,
Qingge Ma^a^,^c ,
Xiangxu Mu^a^,^d ,
Yunzi Luo^a^,^b

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Abstract

Measuring the absolute protein expression quantity for a specific promoter is necessary in the fields of both molecular biology and synthetic biology. The strength of a promoter is traditionally characterized by measuring the fluorescent intensity of the fluorescent protein downstream of the promoter. Until now, measurement of the absolute protein expression quantity for a promoter, however, has been unsuccessful in synthetic biology. The fact that the protein coding sequence influences the expression level for different proteins, and the inconvenience of measuring the absolute protein expression level, present a challenge to absolute quantitative measurement. Here, we introduce a new method that combines the insulator RiboJ with the standard fluorescence curve in order to measure the absolute protein expression quantity quickly; this method has been validated by modeling verification. Using this method, we successfully measured nine constitutive promoters in the Anderson promoter family. Our method provides data with higher accuracy for pathway design and is a straightforward way to standardize the strength of different promoters.

Keywords

RiboJ / Promoter measurement / Synthetic biology

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Hongbin Yu, Zheng Wang, Hanyue Xu, Jiusi Guo, Qingge Ma, Xiangxu Mu, Yunzi Luo. A method for Absolute Protein Expression Quantity Measurement Employing Insulator RiboJ. Engineering, 2018, 4(6): 881‒887 https://doi.org/10.1016/j.eng.2018.09.012

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Acknowledgements

This work was supported by Fundamental Research Funds for Central Universities to Yunzi Luo, the National Natural Science Foundation of China (81502966) and the China Innovation and Entrepreneurship Training Program for Undergraduates (201710611744). We are thankful for the support from our principal investigator (PI) and instructors: Prof. Yunzi Luo, our PI, who provided the lab and supervision necessary for us to finish this project, and Prof. Dan Su, who also provided a lab for us. We thank Huayi Liu and Liping Wang from Yunzi Luo’s lab, and Yiping Chen from Dan Su’s lab of the State Key Lab of Biotherapy for their advice. We also thank other team members from West China Medical Center of Sichuan University for iGEM 2017, and Wen Guo and Yifan Zhong especially. We thank the West China School of Medicine / West China Hospital and the State Key Lab of Biotherapy for their support. We thank the iGEM foundation for providing the opportunity for us to take part in the worldwide biology competition to start our research.