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

Author:

Cepleanu-Pascu Ionut Adrian1ORCID,Stan Miruna2,Cocioba Sebastian3,Stoica Ileana1

Affiliation:

1. Department of Genetics, Faculty of Biology, University of Bucharest, Bucharest, Romania

2. Department of Biochemistry & Molecular Biology, Faculty of Biology, University of Bucharest, Bucharest, Romania

3. Binomica Labs, Long Island City, New York, NY 11101, USA

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.

Publisher

Future Science Ltd

Subject

General Biochemistry, Genetics and Molecular Biology,Biotechnology

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