Easy method for six-fragment Golden Gate Assembly of modular vectors

Abstract

Efficient cloning techniques are a requirement for synthetic biology. This study provides a simplified cloning method based on Golden Gate Assembly that can be used for rapid vector construction. The building of multiple expression vectors with customizable modules is achieved in a timely manner with minimal hands-on time by removing unnecessary steps in the workflow. The authors constructed a total of 21 mammalian episomal expression vectors and conducted a fluorescence expression comparison for different regulatory region combinations post-transfection in HEK293T and HEPG2 cells. Screening revealed that using the EF-1α promoter in combination with the bovine growth hormone polyadenylation sequence seemed to perform best in the types of cells tested compared with other variants.

Keywords

• cloning
• Golden Gate
• Golden Gate standard
• mammalian expression vectors
• S/MARs