Available methods for assembling expression cassettes for synthetic biology |
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Authors: | Tianwen?Wang Email author" target="_blank">Xingyuan?MaEmail author Hu?Zhu Aitao?Li Guocheng?Du Jian?Chen |
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Institution: | (1) State Key Laboratory of Food Science and Technology, Jiangnan University, Wuxi, 214122, Jiangsu, China;(2) School of Biotechnology, and State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 214122, China;(3) Center for Bioengineering and Biotechnology, China University of Petroleum (East China), Qingdao, 266555, China; |
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Abstract: | Studies in the structural biology of the multicomponent protein complex, metabolic engineering, and synthetic biology frequently
rely on the efficient over-expression of these subunits or enzymes in the same cell. As a first step, constructing the multiple
expression cassettes will be a complicated and time-consuming job if the classic and conventional digestion and ligation based
cloning method is used. Some more efficient methods have been developed, including (1) the employment of a multiple compatible
plasmid expression system, (2) the rare-cutter-based design of vectors, (3) in vitro recombination (sequence and ligation
independent cloning, the isothermally enzymatic assembly of DNA molecules in a single reaction), and (4) in vivo recombination
using recombination-efficient yeast (in vivo assembly of overlapping fragments, reiterative recombination for the chromosome
integration of foreign expression cassettes). In this review, we systematically introduce these available methods. |
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