Split intein-mediated selection of cells containing two plasmids using a single antibiotic

Navaneethan Palanisamy, Anna Degen, Anna Morath, Jara Ballestin Ballestin, Claudia Juraske, Mehmet Ali Öztürk, Georg A. Sprenger, Jung-Won Youn, Wolfgang W. Schamel and Barbara Di Ventura ()
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Navaneethan Palanisamy: University of Freiburg
Anna Degen: University of Freiburg
Anna Morath: University of Freiburg
Jara Ballestin Ballestin: University of Freiburg
Claudia Juraske: University of Freiburg
Mehmet Ali Öztürk: University of Freiburg
Georg A. Sprenger: University of Stuttgart
Jung-Won Youn: University of Stuttgart
Wolfgang W. Schamel: University of Freiburg
Barbara Di Ventura: University of Freiburg

Nature Communications, 2019, vol. 10, issue 1, 1-15

Abstract: Abstract To build or dissect complex pathways in bacteria and mammalian cells, it is often necessary to recur to at least two plasmids, for instance harboring orthogonal inducible promoters. Here we present SiMPl, a method based on rationally designed split enzymes and intein-mediated protein trans-splicing, allowing the selection of cells carrying two plasmids with a single antibiotic. We show that, compared to the traditional method based on two antibiotics, SiMPl increases the production of the antimicrobial non-ribosomal peptide indigoidine and the non-proteinogenic aromatic amino acid para-amino-L-phenylalanine from bacteria. Using a human T cell line, we employ SiMPl to obtain a highly pure population of cells double positive for the two chains of the T cell receptor, TCRα and TCRβ, using a single antibiotic. SiMPl has profound implications for metabolic engineering and for constructing complex synthetic circuits in bacteria and mammalian cells.

Date: 2019
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DOI: 10.1038/s41467-019-12911-1

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