Nature Methods
- 4, 705 - 707 (2007)
Published online: 22 July 2007; | doi:10.1038/nmeth1073
High-throughput cloning and expression in recalcitrant bacteriaEric R Geertsma & Bert Poolman
Department of Biochemistry, Groningen Biomolecular Sciences and Biotechnology Institute, and Zernike Institute for Advanced Materials, University of Groningen, Nijenborgh 4, 9747 AG, Groningen, The Netherlands.
Correspondence should be addressed to Bert Poolman b.poolman@rug.nl We developed a generic method for high-throughput cloning in bacteria that are less amenable to conventional DNA manipulations. The method involves ligation-independent cloning in an intermediary Escherichia coli vector, which is rapidly converted via vector-backbone exchange (VBEx) into an organism-specific plasmid ready for high-efficiency transformation. We demonstrated VBEx proof of principle for Lactococcus lactis, but the method can be adapted to all organisms for which plasmids are available.
MORE ARTICLES LIKE THIS These links to content published by NPG are automatically generated.
|
