In vitro DNA SCRaMbLE

Yi Wu, Rui Ying Zhu, Leslie A. Mitchell, Lu Ma, Rui Liu, Meng Zhao, Bin Jia, Hui Xu, Yun Xiang Li, Zu Ming Yang, Yuan Ma, Xia Li, Hong Liu, Duo Liu, Wen Hai Xiao, Xiao Zhou, Bing Zhi Li, Ying Jin Yuan, Jef D. Boeke

    Research output: Contribution to journalArticle

    Abstract

    The power of synthetic biology has enabled the expression of heterologous pathways in cells, as well as genome-scale synthesis projects. The complexity of biological networks makes rational de novo design a grand challenge. Introducing features that confer genetic flexibility is a powerful strategy for downstream engineering. Here we develop an in vitro method of DNA library construction based on structural variation to accomplish this goal. The "in vitro SCRaMbLE system" uses Cre recombinase mixed in a test tube with purified DNA encoding multiple loxPsym sites. Using a β-carotene pathway designed for expression in yeast as an example, we demonstrate top-down and bottom-up in vitro SCRaMbLE, enabling optimization of biosynthetic pathway flux via the rearrangement of relevant transcription units. We show that our system provides a straightforward way to correlate phenotype and genotype and is potentially amenable to biochemical optimization in ways that the in vivo system cannot achieve.

    Original languageEnglish (US)
    Article number3743
    JournalNature communications
    Volume9
    Issue number1
    DOIs
    StatePublished - Dec 1 2018

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    ASJC Scopus subject areas

    • Chemistry(all)
    • Biochemistry, Genetics and Molecular Biology(all)
    • Physics and Astronomy(all)

    Cite this

    Wu, Y., Zhu, R. Y., Mitchell, L. A., Ma, L., Liu, R., Zhao, M., Jia, B., Xu, H., Li, Y. X., Yang, Z. M., Ma, Y., Li, X., Liu, H., Liu, D., Xiao, W. H., Zhou, X., Li, B. Z., Yuan, Y. J., & Boeke, J. D. (2018). In vitro DNA SCRaMbLE. Nature communications, 9(1), [3743]. https://doi.org/10.1038/s41467-018-03743-6