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Split intein-mediated selection of cells containing two plasmids using a single antibiotic
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,...
Autores principales: | , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Nature Publishing Group UK
2019
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6823396/ https://www.ncbi.nlm.nih.gov/pubmed/31672972 http://dx.doi.org/10.1038/s41467-019-12911-1 |
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author | Palanisamy, Navaneethan Degen, Anna Morath, Anna Ballestin Ballestin, Jara Juraske, Claudia Öztürk, Mehmet Ali Sprenger, Georg A. Youn, Jung-Won Schamel, Wolfgang W. Di Ventura, Barbara |
author_facet | Palanisamy, Navaneethan Degen, Anna Morath, Anna Ballestin Ballestin, Jara Juraske, Claudia Öztürk, Mehmet Ali Sprenger, Georg A. Youn, Jung-Won Schamel, Wolfgang W. Di Ventura, Barbara |
author_sort | Palanisamy, Navaneethan |
collection | PubMed |
description | 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. |
format | Online Article Text |
id | pubmed-6823396 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2019 |
publisher | Nature Publishing Group UK |
record_format | MEDLINE/PubMed |
spelling | pubmed-68233962019-11-04 Split intein-mediated selection of cells containing two plasmids using a single antibiotic Palanisamy, Navaneethan Degen, Anna Morath, Anna Ballestin Ballestin, Jara Juraske, Claudia Öztürk, Mehmet Ali Sprenger, Georg A. Youn, Jung-Won Schamel, Wolfgang W. Di Ventura, Barbara Nat Commun Article 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. Nature Publishing Group UK 2019-10-31 /pmc/articles/PMC6823396/ /pubmed/31672972 http://dx.doi.org/10.1038/s41467-019-12911-1 Text en © The Author(s) 2019 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/. |
spellingShingle | Article Palanisamy, Navaneethan Degen, Anna Morath, Anna Ballestin Ballestin, Jara Juraske, Claudia Öztürk, Mehmet Ali Sprenger, Georg A. Youn, Jung-Won Schamel, Wolfgang W. Di Ventura, Barbara Split intein-mediated selection of cells containing two plasmids using a single antibiotic |
title | Split intein-mediated selection of cells containing two plasmids using a single antibiotic |
title_full | Split intein-mediated selection of cells containing two plasmids using a single antibiotic |
title_fullStr | Split intein-mediated selection of cells containing two plasmids using a single antibiotic |
title_full_unstemmed | Split intein-mediated selection of cells containing two plasmids using a single antibiotic |
title_short | Split intein-mediated selection of cells containing two plasmids using a single antibiotic |
title_sort | split intein-mediated selection of cells containing two plasmids using a single antibiotic |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6823396/ https://www.ncbi.nlm.nih.gov/pubmed/31672972 http://dx.doi.org/10.1038/s41467-019-12911-1 |
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