Keywords

Valsa Mali Non-Ribosomal Peptide Synthetase Gene Knockout Complementation

Abstract

[Purpose] Non-ribosomal peptide synthetase (NRPS) plays an important role in the interaction between plant pathogenic fungi and their hosts. Clarifying the function of the VmNRPS12 gene in the pathogenic process of apple tree rot will provide a basis for in-depth research on apple tree rot in the future. Provide theoretical basis for the mechanism of action of bacterial NRPS. [Method] Based on the whole genome data of apple tree rot pathogen, the VmNRPS12 gene was obtained. qRT-PCR technology was used to analyze the expression level of VmNRPS12 in the early stage of infection. Double-joint PCR and PEG-mediated protoplast transformation were used to obtain hygromycin-resistant mutants of the gene. The mutants were detected by PCR and Southern blot verified. Knock out the mutant, and further obtain the complemented mutant by re-introducing the full-length fragment of the gene. Finally, observe the colonies, sporulation and pathogenicity of the wild type, knockout mutant and complemented mutant, and use SPSS software to analyze the detection data. Difference significance analysis. [Results] Quantitative analysis showed that the expression of this gene was significantly up-regulated in the early stage of infection, and the expression level 48 h after inoculation was 138.6 times that of the control. The knockout mutant of this gene has no significant difference in vegetative growth and sporulation compared with the wild-type strain 03-8, but its pathogenicity is significantly weakened compared with the wild-type strain 03-8, and the complemented mutant is pathogenic Forces were restored approximately to wild-type levels. [Conclusion] VmNRPS12 gene is related to the pathogenicity of apple tree rot pathogen

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Original research done by Dai Qingqing, Li Zhengpeng, Han Qingmei, Ma Chenchen, Huang Lili

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