Abstract
Here we report an efficient CRISPR–Cas9 knock-in strategy to activate silent biosynthetic gene clusters (BGCs) in streptomycetes. We applied this one-step strategy to activate multiple BGCs of different classes in five Streptomyces species and triggered the production of unique metabolites, including a novel pentangular type II polyketide in Streptomyces viridochromogenes. This potentially scalable strategy complements existing activation approaches and facilitates discovery efforts to uncover new compounds with interesting bioactivities.
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Acknowledgements
We gratefully acknowledge financial support from the National Institutes of Health (GM077596) (H.Z.), the A*STAR Visiting Investigator Program (H.Z.), and the National Research Foundation, Singapore (NRF2013-THE001-094) (M.M.Z., F.T.W., Y.H.L., E.L.A. and H.Z.). NMR data collection at the UIUC IGB Core was funded by NIH (S10-RR028833). Conjugative donor strains of E. coli were gifted by Prof. William Metcalf at UIUC. We thank members of the Zhao laboratory in UIUC, co-workers in MEL and MERL in A*STAR for constructive comments, Dr. Xudong Guan and Dr. Lingyang Zhu from UIUC for assisting with NMR data acquisition, and Dr. Ying Swan Ho from BTI, A*STAR for HRMS data acquisition.
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M.M.Z., F.T.W., and H.Z. conceived and designed the research. M.M.Z., F.T.W., Y.W., E.H., W.L.Y., R.E.C., and B.E. performed the molecular biology, conjugation and fermentation experiments. S.L. and Y.H.L. performed structure elucidation of compounds. M.M.Z., F.T.W., E.L.A., and H.Z. analyzed the data. M.M.Z., F.T.W., and H.Z. wrote the manuscript.
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Supplementary Results, Supplementary Tables 1–8, Supplementary Figure 1–17 and Supplementary Note (PDF 3726 kb)
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Zhang, M., Wong, F., Wang, Y. et al. CRISPR–Cas9 strategy for activation of silent Streptomyces biosynthetic gene clusters. Nat Chem Biol 13, 607–609 (2017). https://doi.org/10.1038/nchembio.2341
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DOI: https://doi.org/10.1038/nchembio.2341
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