Book chapter · Journal article
Standardized Cloning and Curing of Plasmids
Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark1
Research Groups, Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark2
Microbial Evolution and Synthetic Biology, Research Groups, Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark3
iLoop, Translational Management, Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark4
Bacterial Synthetic Biology, Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark5
Plasmids are highly useful tools for studying living cells and for heterologous expression of genes and pathways in cell factories. Standardized tools and operating procedures for handling such DNA vectors are core principles in synthetic biology. Here, we describe protocols for molecular cloning and exchange of genetic parts in the Standard European Vectors Architecture (SEVA) vector system.
Additionally, to facilitate rapid testing and iterative bioengineering using different vector designs, we provide a one-step protocol for a universal CRISPR-Cas9-based plasmid curing system (pFREE) and demonstrate the application of this system to cure SEVA constructs (all vectors are available at SEVA/Addgene).
| Language: | English |
|---|---|
| Publisher: | Humana Press |
| Year: | 2018 |
| Pages: | 469-476 |
| Series: | Methods in Molecular Biology |
| ISBN: | 1493977946 , 1493977954 , 9781493977949 and 9781493977956 |
| ISSN: | 10643745 and 19406029 |
| Types: | Book chapter and Journal article |
| DOI: | 10.1007/978-1-4939-7795-6_28 |
| ORCIDs: | Lauritsen, Ida , Kim, Se Hyeuk , Porse, Andreas and Nørholm, Morten H. H. |
