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Dissection of the epoxyjanthitrem pathway in Epichloë sp. LpTG-3 strain AR37 by CRISPR gene editing

Epichloë festucae var. lolii and Epichloë sp. LpTG-3 are filamentous fungal endophytes of perennial ryegrass (Lolium perenne) that have a substantial impact on New Zealand’s agricultural economy by conferring biotic advantages to the host grass. Overall, Epichloë endophytes contribute NZ$200 million...

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Autores principales: Miller, Taryn A., Hudson, Debbie A., Johnson, Richard D., Singh, Jaspreet S., Mace, Wade J., Forester, Natasha T., Maclean, Paul H., Voisey, Christine R., Johnson, Linda J.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10512260/
https://www.ncbi.nlm.nih.gov/pubmed/37746172
http://dx.doi.org/10.3389/ffunb.2022.944234
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author Miller, Taryn A.
Hudson, Debbie A.
Johnson, Richard D.
Singh, Jaspreet S.
Mace, Wade J.
Forester, Natasha T.
Maclean, Paul H.
Voisey, Christine R.
Johnson, Linda J.
author_facet Miller, Taryn A.
Hudson, Debbie A.
Johnson, Richard D.
Singh, Jaspreet S.
Mace, Wade J.
Forester, Natasha T.
Maclean, Paul H.
Voisey, Christine R.
Johnson, Linda J.
author_sort Miller, Taryn A.
collection PubMed
description Epichloë festucae var. lolii and Epichloë sp. LpTG-3 are filamentous fungal endophytes of perennial ryegrass (Lolium perenne) that have a substantial impact on New Zealand’s agricultural economy by conferring biotic advantages to the host grass. Overall, Epichloë endophytes contribute NZ$200 million to the economy annually, with strain AR37 estimated to contribute NZ$3.6 billion to the New Zealand economy over a 20-year period. This strain produces secondary metabolites, including epoxyjanthitrems, which are a class of indole diterpenes, associated with the observed effects of AR37 on livestock and insect pests. Until very recently, AR37 was intractable to genetic modification but this has changed with the application of CRISPR-Cas9 based gene editing techniques. In this paper, gene inactivation by CRISPR-Cas9 was used to deconvolute the genetic basis for epoxyjanthitrem biosynthesis, including creating an AR37 strain that has been edited to remove the biosynthesis of all indole diterpenes. We show that gene editing of Epichloë can be achieved without off-target events or introduction of foreign DNA (footprint-less) through an AMA1-based plasmid that simultaneously expresses the CRISPR-Cas9 system and selectable marker. Genetic modification events in these transformants were investigated through genome sequencing and in planta chemistry.
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spelling pubmed-105122602023-09-22 Dissection of the epoxyjanthitrem pathway in Epichloë sp. LpTG-3 strain AR37 by CRISPR gene editing Miller, Taryn A. Hudson, Debbie A. Johnson, Richard D. Singh, Jaspreet S. Mace, Wade J. Forester, Natasha T. Maclean, Paul H. Voisey, Christine R. Johnson, Linda J. Front Fungal Biol Fungal Biology Epichloë festucae var. lolii and Epichloë sp. LpTG-3 are filamentous fungal endophytes of perennial ryegrass (Lolium perenne) that have a substantial impact on New Zealand’s agricultural economy by conferring biotic advantages to the host grass. Overall, Epichloë endophytes contribute NZ$200 million to the economy annually, with strain AR37 estimated to contribute NZ$3.6 billion to the New Zealand economy over a 20-year period. This strain produces secondary metabolites, including epoxyjanthitrems, which are a class of indole diterpenes, associated with the observed effects of AR37 on livestock and insect pests. Until very recently, AR37 was intractable to genetic modification but this has changed with the application of CRISPR-Cas9 based gene editing techniques. In this paper, gene inactivation by CRISPR-Cas9 was used to deconvolute the genetic basis for epoxyjanthitrem biosynthesis, including creating an AR37 strain that has been edited to remove the biosynthesis of all indole diterpenes. We show that gene editing of Epichloë can be achieved without off-target events or introduction of foreign DNA (footprint-less) through an AMA1-based plasmid that simultaneously expresses the CRISPR-Cas9 system and selectable marker. Genetic modification events in these transformants were investigated through genome sequencing and in planta chemistry. Frontiers Media S.A. 2022-08-10 /pmc/articles/PMC10512260/ /pubmed/37746172 http://dx.doi.org/10.3389/ffunb.2022.944234 Text en Copyright © 2022 Miller, Hudson, Johnson, Singh, Mace, Forester, Maclean, Voisey and Johnson https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Fungal Biology
Miller, Taryn A.
Hudson, Debbie A.
Johnson, Richard D.
Singh, Jaspreet S.
Mace, Wade J.
Forester, Natasha T.
Maclean, Paul H.
Voisey, Christine R.
Johnson, Linda J.
Dissection of the epoxyjanthitrem pathway in Epichloë sp. LpTG-3 strain AR37 by CRISPR gene editing
title Dissection of the epoxyjanthitrem pathway in Epichloë sp. LpTG-3 strain AR37 by CRISPR gene editing
title_full Dissection of the epoxyjanthitrem pathway in Epichloë sp. LpTG-3 strain AR37 by CRISPR gene editing
title_fullStr Dissection of the epoxyjanthitrem pathway in Epichloë sp. LpTG-3 strain AR37 by CRISPR gene editing
title_full_unstemmed Dissection of the epoxyjanthitrem pathway in Epichloë sp. LpTG-3 strain AR37 by CRISPR gene editing
title_short Dissection of the epoxyjanthitrem pathway in Epichloë sp. LpTG-3 strain AR37 by CRISPR gene editing
title_sort dissection of the epoxyjanthitrem pathway in epichloë sp. lptg-3 strain ar37 by crispr gene editing
topic Fungal Biology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10512260/
https://www.ncbi.nlm.nih.gov/pubmed/37746172
http://dx.doi.org/10.3389/ffunb.2022.944234
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