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CRISPR/Cas9‐RNA interference system for combinatorial metabolic engineering of Saccharomyces cerevisiae

The yeast Saccharomyces cerevisiae is widely used in industrial biotechnology for the production of fuels, chemicals, food ingredients, food and beverages, and pharmaceuticals. To obtain high‐performing strains for such bioprocesses, it is often necessary to test tens or even hundreds of metabolic e...

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Autores principales: Kildegaard, Kanchana Rueksomtawin, Tramontin, Larissa Ribeiro Ramos, Chekina, Ksenia, Li, Mingji, Goedecke, Tobias Justus, Kristensen, Mette, Borodina, Irina
Formato: Online Artículo Texto
Lenguaje:English
Publicado: John Wiley and Sons Inc. 2019
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6619288/
https://www.ncbi.nlm.nih.gov/pubmed/30953378
http://dx.doi.org/10.1002/yea.3390
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author Kildegaard, Kanchana Rueksomtawin
Tramontin, Larissa Ribeiro Ramos
Chekina, Ksenia
Li, Mingji
Goedecke, Tobias Justus
Kristensen, Mette
Borodina, Irina
author_facet Kildegaard, Kanchana Rueksomtawin
Tramontin, Larissa Ribeiro Ramos
Chekina, Ksenia
Li, Mingji
Goedecke, Tobias Justus
Kristensen, Mette
Borodina, Irina
author_sort Kildegaard, Kanchana Rueksomtawin
collection PubMed
description The yeast Saccharomyces cerevisiae is widely used in industrial biotechnology for the production of fuels, chemicals, food ingredients, food and beverages, and pharmaceuticals. To obtain high‐performing strains for such bioprocesses, it is often necessary to test tens or even hundreds of metabolic engineering targets, preferably in combinations, to account for synergistic and antagonistic effects. Here, we present a method that allows simultaneous perturbation of multiple selected genetic targets by combining the advantage of CRISPR/Cas9, in vivo recombination, USER assembly and RNA interference. CRISPR/Cas9 introduces a double‐strand break in a specific genomic region, where multiexpression constructs combined with the knockdown constructs are simultaneously integrated by homologous recombination. We show the applicability of the method by improving cis,cis‐muconic acid production in S. cerevisiae through simultaneous manipulation of several metabolic engineering targets. The method can accelerate metabolic engineering efforts for the construction of future cell factories.
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spelling pubmed-66192882019-07-22 CRISPR/Cas9‐RNA interference system for combinatorial metabolic engineering of Saccharomyces cerevisiae Kildegaard, Kanchana Rueksomtawin Tramontin, Larissa Ribeiro Ramos Chekina, Ksenia Li, Mingji Goedecke, Tobias Justus Kristensen, Mette Borodina, Irina Yeast Special Issue Articles The yeast Saccharomyces cerevisiae is widely used in industrial biotechnology for the production of fuels, chemicals, food ingredients, food and beverages, and pharmaceuticals. To obtain high‐performing strains for such bioprocesses, it is often necessary to test tens or even hundreds of metabolic engineering targets, preferably in combinations, to account for synergistic and antagonistic effects. Here, we present a method that allows simultaneous perturbation of multiple selected genetic targets by combining the advantage of CRISPR/Cas9, in vivo recombination, USER assembly and RNA interference. CRISPR/Cas9 introduces a double‐strand break in a specific genomic region, where multiexpression constructs combined with the knockdown constructs are simultaneously integrated by homologous recombination. We show the applicability of the method by improving cis,cis‐muconic acid production in S. cerevisiae through simultaneous manipulation of several metabolic engineering targets. The method can accelerate metabolic engineering efforts for the construction of future cell factories. John Wiley and Sons Inc. 2019-06-13 2019-05 /pmc/articles/PMC6619288/ /pubmed/30953378 http://dx.doi.org/10.1002/yea.3390 Text en © 2019 The Authors Yeast Published by John Wiley & Sons Ltd This is an open access article under the terms of the http://creativecommons.org/licenses/by/4.0/ License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.
spellingShingle Special Issue Articles
Kildegaard, Kanchana Rueksomtawin
Tramontin, Larissa Ribeiro Ramos
Chekina, Ksenia
Li, Mingji
Goedecke, Tobias Justus
Kristensen, Mette
Borodina, Irina
CRISPR/Cas9‐RNA interference system for combinatorial metabolic engineering of Saccharomyces cerevisiae
title CRISPR/Cas9‐RNA interference system for combinatorial metabolic engineering of Saccharomyces cerevisiae
title_full CRISPR/Cas9‐RNA interference system for combinatorial metabolic engineering of Saccharomyces cerevisiae
title_fullStr CRISPR/Cas9‐RNA interference system for combinatorial metabolic engineering of Saccharomyces cerevisiae
title_full_unstemmed CRISPR/Cas9‐RNA interference system for combinatorial metabolic engineering of Saccharomyces cerevisiae
title_short CRISPR/Cas9‐RNA interference system for combinatorial metabolic engineering of Saccharomyces cerevisiae
title_sort crispr/cas9‐rna interference system for combinatorial metabolic engineering of saccharomyces cerevisiae
topic Special Issue Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6619288/
https://www.ncbi.nlm.nih.gov/pubmed/30953378
http://dx.doi.org/10.1002/yea.3390
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