Leptosphaeria maculans is the causal agent of blackleg disease on Brassica napus. Determining the underlying functions of genes required for pathogenesis is essential for understanding the infection process. A chitin-binding protein (LmCBP1) was discovered as a pathogenicity factor for the infection of B. napus by L. maculans through gene knockout using the CRISPR-Cas9 system. Chitin-binding activity was demonstrated through a chitin-protein binding assay. A secreted signal peptide was detected using a yeast secreted-signal peptide trap assay. An increased expression level during the infection stage was also observed, suggesting that LmCBP1 is a secreted protein. The knockout mutants showed decreased infection on B. napus, with reduced pathogenicity on ten cultivars with/without diverse R genes. The mutants were more sensitive to H2O2 compared to wild type L. maculans isolate JN3. This study provides evidence of the virulence of a novel chitin-binding protein LmCBP1 on B. napus through mutants created via the CRISPR-Cas9 system.

中文翻译：

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LmCBP1是一种分泌的几丁质结合蛋白，对于油菜对斑节菜的致病力是必需的。

Leptosphaeria maculans是甘蓝型油菜上黑腿病的病原体。确定发病机理所需基因的基本功能对于理解感染过程至关重要。发现使用几丁质结合蛋白（LmCBP1）作为致病因子，可通过使用CRISPR-Cas9系统进行基因敲除来感染黄斑狼疮。通过几丁质蛋白结合测定法证明了几丁质结合活性。使用酵母分泌信号肽捕获测定法检测了分泌信号肽。在感染阶段还观察到表达水平升高，表明LmCBP1是一种分泌蛋白。敲除突变体显示对甘蓝型油菜的感染减少，对具有/不具有多种R基因的十个品种的致病性降低。与野生型黄斑狼疮分离株JN3相比，该突变体对H2O2更为敏感。这项研究提供了一种新的几丁质结合蛋白LmCBP1通过CRISPR-Cas9系统产生的突变体对油菜的致病力的证据。