利用纳米磁珠介导的茄链格孢菌遗传转化技术的建立及其泛素化蛋白酶基因<em>AsPR45</em>功能研究

doi:10.3969/j.issn.1674-7968.2025.09.014

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Abstract The ubiquitin-proteasome system (UPS) is a critical protein degradation pathway in fungal cells, playing a vital role in regulating protein homeostasis and maintaining cellular functions. Previous studies have shown that during the early stages (0~72 h) of Alternaria solani infection in potato (Solanum tuberosum), the expression of the AsPR45 gene, which encoded the P45 subunit of the 26S proteasome in A. solani, was significantly upregulated. To further explore the function of the AsPR45 gene, this study first optimized a genetic transformation method for A. solani mediated by magnetic nanoparticles (MNPs). Using this transformation assay, a premature termination construct of AsPR45 was developed based on the pTOR-mRFP vector backbone and introduced into the wild-type strain TA-0410. Functional validation was carried out on 2 positive mutant strains. The transformation efficiency was the highest when the mass ratio of MNPs to the pTOR-mRFP vector was 1∶4, with binding at 37 ℃ for 30 min, followed by co-incubation with spores at room temperature for 1.5 h, achieving a transformation rate of (36.77±2.91)%. Biological function assays revealed that premature termination mutants of AsPR45 exhibited no significant difference in hyphal growth rate compared with the wild-type strain. However, the mutants produced less melanin in hyphae, had a significantly reduced conidiation rate, and showed markedly decreased pathogenicity on the leaves of potato cultivar 'Desiree'. Additionally, the EC₅₀ values for difenoconazole resistance differed significantly between the wild-type strain and the mutants. These results indicated that AsPR45 was a key gene in the ubiquitination pathway in A. solani, playing an important role in fungal morphogenesis, potato infection, and stress resistance. This study not only provides a technical foundation for functional gene research in A. solani but also contributes to the identification of potential fungicide targets, accelerating the development of strategies for controlling potato early blight disease.

Key words： Alternaria solani Nanoparticles Transformation Ubiquitin-proteasome pathway Mutant

Received: 21 October 2024

ZTFLH:

S435.32

Corresponding Authors: ^**4311@ynnu.edu.cn; lijianmei152927@163.com

About author:: ^*These authors contributed equally to this work

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	ZHAO Yu-Jiao
	FU Ai-Hua
	ZHANG Jia-Ying
	LIU Wen-Wen
	YANG Qiao-Mei
	JIANG Dong-Mei
	TANG Wei
	LI Jian-Mei

Cite this article:

ZHAO Yu-Jiao,FU Ai-Hua,ZHANG Jia-Ying, et al. Development of a Magnetic Nanoparticle-mediated Genetic Transformation Technology for Alternaria solani and Functional Analysis of the Ubiquitin-proteasome Gene AsPR45[J]. 农业生物技术学报, 2025, 33(9): 2036-2048.

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https://journal05.magtech.org.cn/Jwk_ny/EN/10.3969/j.issn.1674-7968.2025.09.014 OR https://journal05.magtech.org.cn/Jwk_ny/EN/Y2025/V33/I9/2036

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