In this section
@article{Moore:2021:10.1021/acssynbio.0c00581,
author = {Moore, SJ and Lai, H-E and Chee, S-M and Toh, M and Coode, S and Chengan, K and Capel, P and Corre, C and de, los Santos ELC and Freemont, PS},
doi = {10.1021/acssynbio.0c00581},
journal = {ACS Synthetic Biology},
pages = {402--411},
title = {A Streptomyces venezuelae cell-free toolkit for synthetic biology},
url = {http://dx.doi.org/10.1021/acssynbio.0c00581},
volume = {10},
year = {2021}
}
TY - JOUR
AB - Prokaryotic cell-free coupled transcription–translation (TX-TL) systems are emerging as a powerful tool to examine natural product biosynthetic pathways in a test tube. The key advantages of this approach are the reduced experimental time scales and controlled reaction conditions. To realize this potential, it is essential to develop specialized cell-free systems in organisms enriched for biosynthetic gene clusters. This requires strong protein production and well-characterized synthetic biology tools. The Streptomyces genus is a major source of natural products. To study enzymes and pathways from Streptomyces, we originally developed a homologous Streptomyces cell-free system to provide a native protein folding environment, a high G+C (%) tRNA pool, and an active background metabolism. However, our initial yields were low (36 μg/mL) and showed a high level of batch-to-batch variation. Here, we present an updated high-yield and robust Streptomyces TX-TL protocol, reaching up to yields of 266 μg/mL of expressed recombinant protein. To complement this, we rapidly characterize a range of DNA parts with different reporters, express high G+C (%) biosynthetic genes, and demonstrate an initial proof of concept for combined transcription, translation, and biosynthesis of Streptomyces metabolic pathways in a single “one-pot” reaction.
AU - Moore,SJ
AU - Lai,H-E
AU - Chee,S-M
AU - Toh,M
AU - Coode,S
AU - Chengan,K
AU - Capel,P
AU - Corre,C
AU - de,los Santos ELC
AU - Freemont,PS
DO - 10.1021/acssynbio.0c00581
EP - 411
PY - 2021///
SN - 2161-5063
SP - 402
TI - A Streptomyces venezuelae cell-free toolkit for synthetic biology
T2 - ACS Synthetic Biology
UR - http://dx.doi.org/10.1021/acssynbio.0c00581
UR - https://www.webofscience.com/api/gateway?GWVersion=2&SrcApp=PARTNER_APP&SrcAuth=LinksAMR&KeyUT=WOS:000621601500019&DestLinkType=FullRecord&DestApp=ALL_WOS&UsrCustomerID=a2bf6146997ec60c407a63945d4e92bb
VL - 10
ER -
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