Michael Smith Laboratories, University of British Columbia, Vancouver, BC V6T 1Z4, Canada.
Department of Botany, University of British Columbia, Vancouver, BC V6T 1Z4, Canada.
Mol Plant Microbe Interact. 2022 Mar;35(3):244-256. doi: 10.1094/MPMI-10-21-0254-R. Epub 2022 Mar 10.
Most plant fungal pathogens that cause worldwide crop losses are understudied, due to various technical challenges. With the increasing availability of sequenced whole genomes of these non-model fungi, effective genetic analysis methods are highly desirable. Here, we describe a newly developed pipeline, which combines forward genetic screening with high-throughput next-generation sequencing to enable quick gene discovery. We applied this pipeline in the notorious soilborne phytopathogen and identified 32 mutants with various developmental and growth deficiencies. Detailed molecular studies of three melanization-deficient mutants provide a proof of concept for the effectiveness of our method. A master transcription factor was found to regulate melanization of sclerotia through the DHN (1,8-dihydroxynaphthalene) melanin biosynthesis pathway. In addition, these mutants revealed that sclerotial melanization is important for sclerotia survival under abiotic stresses, sclerotial surface structure, and sexual reproduction. Foreseeably, this pipeline can be applied to facilitate efficient in-depth studies of other non-model fungal species in the future.[Formula: see text] Copyright © 2022 The Author(s). This is an open access article distributed under the CC BY 4.0 International license.
由于各种技术挑战，大多数导致全球作物损失的植物病原真菌研究不足。随着这些非模式真菌测序全基因组的日益普及，人们非常希望开发有效的遗传分析方法。在这里，我们描述了一种新开发的流水线，它将正向遗传筛选与高通量下一代测序相结合，从而实现快速基因发现。我们将该流水线应用于臭名昭著的土传植物病原菌，并鉴定出 32 种具有各种发育和生长缺陷的突变体。对三种黑色素缺陷突变体的详细分子研究为我们方法的有效性提供了概念验证。发现一个主转录因子通过 DHN（1,8-二羟基萘）黑色素生物合成途径调节菌核的黑色素化。此外，这些突变体表明菌核黑色素化对于非生物胁迫下菌核的存活、菌核表面结构和有性生殖很重要。可以预见，该流水线将来可用于促进对其他非模式真菌物种的高效深入研究。[公式：见正文] 版权所有© 2022 作者。这是一个在 CC BY 4.0 国际许可下发布的开放获取文章。
