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Objective To explore the functions of type XI histidine kinase genes in Botrytis cinerea, thereby establishing a foundation for further elucidation of the molecular mechanisms involving histidine kinase-related genes in the growth, development, and pathogenic processes of plant pathogenic fungi. Methods Through knockout vector construction, ATMT transformation, and PCR/qPCR validation, we obtained the mutant ΔBcHK91. The colony growth, sclerotium formation, conidial production and morphology, conidial germination rate, appressorium and infection cushion formation rates, pathogenicity, cell wall/membrane integrity assays and transcriptome profiling were conducted to evaluate the effect of BcHK91 knockout on B. cinerea strain B05.10, thereby elucidating the biological function of this gene. Results Two BcHK91 knockout mutants were successfully obtained and designated as ΔBcHK91-A and ΔBcHK91-B. The phenotypic analysis revealed that the knockout of BcHK91 reduced the conidial length, conidial production, conidial germination rate, appressorium formation rate, infection cushion formation rate, and pathogenicity. Moreover, the mutant formed no sclerotium and showed increased sensitivityto Congo red and SDS compared with the wild-type strain B05.10 and the ectopic insertion strain ET. To reveal the transcription mechanisms of BcHK91, we compared the transcriptomes of B05.10 and ΔBcHK91 by RNA-seq. A total of 1 533 differentially expressed genes (DEGs) were predicted in ΔBcHK91, including 1 017 genes with up-regulated expression and 516 genes with down-regulated expression. Gene ontology (GO) and clusters of orthologous groups (COG) annotation showed that the DEGs were mainly involved in cell process and metabolic process of biological processes, cellular anatomical entity and intracellular of cellular components, and catalytic activity, binding, and transporter activity of molecular functions. The Kyoto encyclopedia of genes and genomes (KEGG) pathway analysis showed that the DEGs were mainly involved in carbohydrate transport and metabolism, starch and sucrose metabolism, and mitogen-activated protein kinase (MAPK) cascade response and other physiological metabolic pathways related tothe phenotype and pathogenicity of ΔBcHK91. In silico analysis of the DEGs suggested that 17 DEGs were related to the growth and development, oxidative stress, cell wall biosynthesis, cell membrane integrity, sclerotial initials, and pathogenicity. The results of qPCR demonstrated that the expression levels of 13 genes matched the trends observed in RNA-seq data, confirming the high reliability of the transcriptome analysis. Conclusion In B. cinerea, BcHK91 plays critical roles in asexual development, environmental stress responses, and pathogenicity.
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| 科 Family | 属数 Number of genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) | 属 Genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) |
|---|---|---|---|---|---|---|
| 鹅膏菌科Amanitaceae | 2 | 11 | 5.26 | 鹅膏菌属 Amanita | 10 | 4.78 |
| 小菇科 Mycenaceae | 2 | 12 | 5.74 | 丝盖伞属 Inocybe | 5 | 2.39 |
| 多孔菌科 Polyporaceae | 8 | 14 | 6.70 | 蜡蘑属 Laccaria | 5 | 2.39 |
| 红菇科 Russulaceae | 3 | 23 | 11.00 | 小皮伞属 Marasmius | 6 | 2.87 |
| 小菇属 Mycena | 11 | 5.26 | ||||
| 光柄菇属 Pluteus | 5 | 2.39 | ||||
| 红菇属 Russula | 17 | 8.13 | ||||
| 栓菌属 Trametes | 5 | 2.39 |
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