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

Autor:	Miller TA; Grasslands Research Centre, AgResearch, Palmerston North, New Zealand., Hudson DA; Grasslands Research Centre, AgResearch, Palmerston North, New Zealand., Johnson RD; Grasslands Research Centre, AgResearch, Palmerston North, New Zealand., Singh JS; Grasslands Research Centre, AgResearch, Palmerston North, New Zealand., Mace WJ; Grasslands Research Centre, AgResearch, Palmerston North, New Zealand., Forester NT; Grasslands Research Centre, AgResearch, Palmerston North, New Zealand., Maclean PH; Grasslands Research Centre, AgResearch, Palmerston North, New Zealand., Voisey CR; Grasslands Research Centre, AgResearch, Palmerston North, New Zealand., Johnson LJ; Grasslands Research Centre, AgResearch, Palmerston North, New Zealand.
Jazyk:	angličtina
Zdroj:	Frontiers in fungal biology [Front Fungal Biol] 2022 Aug 10; Vol. 3, pp. 944234. Date of Electronic Publication: 2022 Aug 10 (Print Publication: 2022).
DOI:	10.3389/ffunb.2022.944234
Abstrakt:	Epichloë festucae var. lolii and Epichloë sp. Lp TG-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. Competing Interests: The authors declare that this study received funding from Grasslanz Technology Ltd and PGG Wrightson Seeds Ltd. The funders were not involved in the study design, collection, analysis, interpretation of dat or the writing of this article. They were however involved in the decision to submit it for publication. (Copyright © 2022 Miller, Hudson, Johnson, Singh, Mace, Forester, Maclean, Voisey and Johnson.)
Databáze:	MEDLINE
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