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Biotechnol J. 2017 Apr;12(4). doi: 10.1002/biot.201600678. Epub 2017 Feb 24.

Streptomyces venezuelae TX-TL - a next generation cell-free synthetic biology tool.

Author information

1
Centre for Synthetic Biology and Innovation, South Kensington Campus, London, UK.
2
Department of Medicine, South Kensington Campus, London, UK.
3
Department of Life Science, South Kensington Campus, London, UK.

Abstract

Streptomyces venezuelae is a promising chassis in synthetic biology for fine chemical and secondary metabolite pathway engineering. The potential of S. venezuelae could be further realized by expanding its capability with the introduction of its own in vitro transcription-translation (TX-TL) system. TX-TL is a fast and expanding technology for bottom-up design of complex gene expression tools, biosensors and protein manufacturing. Herein, we introduce a S. venezuelae TX-TL platform by reporting a streamlined protocol for cell-extract preparation, demonstrating high-yield synthesis of a codon-optimized sfGFP reporter and the prototyping of a synthetic tetracycline-inducible promoter in S. venezuelae TX-TL based on the tetO-TetR repressor system. The aim of this system is to provide a host for the homologous production of exotic enzymes from Actinobacteria secondary metabolism in vitro. As an example, the authors demonstrate the soluble synthesis of a selection of enzymes (12-70 kDa) from the Streptomyces rimosus oxytetracycline pathway.

KEYWORDS:

Cell-free; Gene expression; Streptomyces; Synthetic biology; Tetracycline

PMID:
28139884
DOI:
10.1002/biot.201600678
[Indexed for MEDLINE]
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