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Model-guided engineering of DNA sequences with predictable site-specific recombination rates

Author

Listed:
  • Qiuge Zhang

    (University of Minnesota)

  • Samira M. Azarin

    (University of Minnesota)

  • Casim A. Sarkar

    (University of Minnesota)

Abstract

Site-specific recombination (SSR) is an important tool in synthetic biology, but its applications are limited by the inability to predictably tune SSR reaction rates. Facile rate manipulation could be achieved by modifying the DNA substrate sequence; however, this approach lacks rational design principles. Here, we develop an integrated experimental and computational method to engineer the DNA attachment sequence attP for predictably modulating the inversion reaction mediated by the recombinase Bxb1. After developing a qPCR method to measure SSR reaction rate, we design, select, and sequence attP libraries to inform a machine-learning model that computes Bxb1 inversion rate as a function of attP sequence. We use this model to predict reaction rates of attP variants in vitro and demonstrate their utility in gene circuit design in Escherichia coli. Our high-throughput, model-guided approach for rationally tuning SSR reaction rates enhances our understanding of recombinase function and expands the synthetic biology toolbox.

Suggested Citation

  • Qiuge Zhang & Samira M. Azarin & Casim A. Sarkar, 2022. "Model-guided engineering of DNA sequences with predictable site-specific recombination rates," Nature Communications, Nature, vol. 13(1), pages 1-12, December.
    • Handle: RePEc:nat:natcom:v:13:y:2022:i:1:d:10.1038_s41467-022-31538-3
    DOI: 10.1038/s41467-022-31538-3
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    References listed on IDEAS

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    1. U. T. Bornscheuer & G. W. Huisman & R. J. Kazlauskas & S. Lutz & J. C. Moore & K. Robins, 2012. "Engineering the third wave of biocatalysis," Nature, Nature, vol. 485(7397), pages 185-194, May.
    2. Ming-Ru Wu & Lior Nissim & Doron Stupp & Erez Pery & Adina Binder-Nissim & Karen Weisinger & Casper Enghuus & Sebastian R. Palacios & Melissa Humphrey & Zhizhuo Zhang & Eva Maria Novoa & Manolis Kelli, 2019. "A high-throughput screening and computation platform for identifying synthetic promoters with enhanced cell-state specificity (SPECS)," Nature Communications, Nature, vol. 10(1), pages 1-10, December.
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