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Gene manipulation in the Mucorales fungus Rhizopus oryzae using TALENs with exonuclease overexpression

The Mucorales fungal genus Rhizopus is used for the industrial production of organic acids, enzymes and fermented foods. The metabolic engineering efficiency of Rhizopus could be improved using gene manipulation; however, exogenous DNA rarely integrates into the host genome. Consequently, a genetic...

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Autores principales: Tsuboi, Yuichi, Sakuma, Tetsushi, Yamamoto, Takashi, Horiuchi, Hiroyuki, Takahashi, Fumikazu, Igarashi, Kazuaki, Hagihara, Hiroshi, Takimura, Yasushi
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Oxford University Press 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8863565/
https://www.ncbi.nlm.nih.gov/pubmed/35137045
http://dx.doi.org/10.1093/femsle/fnac010
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author Tsuboi, Yuichi
Sakuma, Tetsushi
Yamamoto, Takashi
Horiuchi, Hiroyuki
Takahashi, Fumikazu
Igarashi, Kazuaki
Hagihara, Hiroshi
Takimura, Yasushi
author_facet Tsuboi, Yuichi
Sakuma, Tetsushi
Yamamoto, Takashi
Horiuchi, Hiroyuki
Takahashi, Fumikazu
Igarashi, Kazuaki
Hagihara, Hiroshi
Takimura, Yasushi
author_sort Tsuboi, Yuichi
collection PubMed
description The Mucorales fungal genus Rhizopus is used for the industrial production of organic acids, enzymes and fermented foods. The metabolic engineering efficiency of Rhizopus could be improved using gene manipulation; however, exogenous DNA rarely integrates into the host genome. Consequently, a genetic tool for Mucorales fungi needs to be developed. Recently, programmable nucleases that generate DNA double-strand breaks (DSBs) at specific genomic loci have been used for genome editing in various organisms. In this study, we examined gene disruption in Rhizopus oryzae using transcription activator-like effector nucleases (TALENs), with and without exonuclease overexpression. TALENs with an overexpressing exonuclease induced DSBs, followed by target site deletions. Although DSBs are repaired mainly by nonhomologous end joining in most organisms, our results suggested that in R. oryzae microhomology-mediated end joining was the major DSB repair system. Our gene manipulation method using TALENs coupled with exonuclease overexpression contributes to basic scientific knowledge and the metabolic engineering of Rhizopus.
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spelling pubmed-88635652022-02-23 Gene manipulation in the Mucorales fungus Rhizopus oryzae using TALENs with exonuclease overexpression Tsuboi, Yuichi Sakuma, Tetsushi Yamamoto, Takashi Horiuchi, Hiroyuki Takahashi, Fumikazu Igarashi, Kazuaki Hagihara, Hiroshi Takimura, Yasushi FEMS Microbiol Lett Research Letter The Mucorales fungal genus Rhizopus is used for the industrial production of organic acids, enzymes and fermented foods. The metabolic engineering efficiency of Rhizopus could be improved using gene manipulation; however, exogenous DNA rarely integrates into the host genome. Consequently, a genetic tool for Mucorales fungi needs to be developed. Recently, programmable nucleases that generate DNA double-strand breaks (DSBs) at specific genomic loci have been used for genome editing in various organisms. In this study, we examined gene disruption in Rhizopus oryzae using transcription activator-like effector nucleases (TALENs), with and without exonuclease overexpression. TALENs with an overexpressing exonuclease induced DSBs, followed by target site deletions. Although DSBs are repaired mainly by nonhomologous end joining in most organisms, our results suggested that in R. oryzae microhomology-mediated end joining was the major DSB repair system. Our gene manipulation method using TALENs coupled with exonuclease overexpression contributes to basic scientific knowledge and the metabolic engineering of Rhizopus. Oxford University Press 2022-02-08 /pmc/articles/PMC8863565/ /pubmed/35137045 http://dx.doi.org/10.1093/femsle/fnac010 Text en © The Author(s) 2022. Published by Oxford University Press on behalf of FEMS. https://creativecommons.org/licenses/by-nc/4.0/This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (https://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com
spellingShingle Research Letter
Tsuboi, Yuichi
Sakuma, Tetsushi
Yamamoto, Takashi
Horiuchi, Hiroyuki
Takahashi, Fumikazu
Igarashi, Kazuaki
Hagihara, Hiroshi
Takimura, Yasushi
Gene manipulation in the Mucorales fungus Rhizopus oryzae using TALENs with exonuclease overexpression
title Gene manipulation in the Mucorales fungus Rhizopus oryzae using TALENs with exonuclease overexpression
title_full Gene manipulation in the Mucorales fungus Rhizopus oryzae using TALENs with exonuclease overexpression
title_fullStr Gene manipulation in the Mucorales fungus Rhizopus oryzae using TALENs with exonuclease overexpression
title_full_unstemmed Gene manipulation in the Mucorales fungus Rhizopus oryzae using TALENs with exonuclease overexpression
title_short Gene manipulation in the Mucorales fungus Rhizopus oryzae using TALENs with exonuclease overexpression
title_sort gene manipulation in the mucorales fungus rhizopus oryzae using talens with exonuclease overexpression
topic Research Letter
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8863565/
https://www.ncbi.nlm.nih.gov/pubmed/35137045
http://dx.doi.org/10.1093/femsle/fnac010
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