CRISPR/Cas9-mediated deletion of large chromosomal segments identifies a minichromosome modulating the Colletotrichum graminicola virulence on maize

文献类型： 外文期刊

第一作者： Ma, Wendi

作者： Ma, Wendi;Yang, Jun;Zhao, Wensheng;Peng, You-Liang;Bhadauria, Vijai;Yang, Jun;Zhao, Wensheng;Peng, You-Liang;Bhadauria, Vijai;Ding, Junqiang;Duan, Canxing

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关键词： CRISPR; Cas9; Anthracnose leaf blight; Anthracnose stem rot; Dispensable chromosomes; Genome editing; Fungal virulence and pathogenicity

期刊名称：INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES （ 影响因子：8.2； 五年影响因子：7.8 ）

ISSN： 0141-8130

年卷期： 2023 年 245 卷

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收录情况： SCI

摘要： Colletotrichum graminicola causes anthracnose on maize, an economically significant disease worldwide. To decipher how the pathogen controls its virulence/pathogenicity on maize at the minichromosomal level, we sequenced the genome and transcriptome of the C. graminicola strain T1-3-3. The 61.91 Mb genome contains three transcriptionally repressed, full-length strain-specific minichromosomes (<1 Mb; Chr11 through Chr13). A CRISPR/Cas9-based system was developed to knock out large chromosomal segments; it involved the generation of multiple simultaneous DNA double-strand breaks across a targeted genomic region, followed by homology -directed replacement thereof with a donor DNA template carrying the selectable marker hygromycin phospho-transferase gene flanked by homologous sequence arms of the targeted region. Using this system, we obtained distinct mutants functionally nullisomic for individual minichromosomes. Only the & UDelta;Chr12 mutant lacking the 498.44 Kb genomic region carrying all of the 31 genes of Chr12 exhibited attenuated virulence on maize and was indistinguishable from T1-3-3 in fungal growth and conidiation, indicating that Chr12 is a conditionally dispensable minichromosome and imparts full virulence to C. graminicola on maize. The CRISPR/Cas9-mediated genome editing system developed in this study will enable the determination of the biological functions of minichromosomes or large chromosomal segments in fungal plant pathogens.

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