Function of nonribosomal peptide synthetase gene VmNRPS12 of Valsa mali

Abstract:

[Objective] Nonribosomal peptide synthetase (NRPS) plays an important role in plant-pathogenic fungi interaction. In this study, we analyzed the role of VmNRPS12 during Valsa mali (V. mali) infection and pathogenecity, which may shed new insights into the pathogenesis of V. mali.[Methods] Based on the genome of V. mali, we obtained one NPRS, designated as VmNRPS12. The expression profile of VmNRPS12 was analyzed by qRT-PCR. Through Double-joint PCR and PEG-mediated protoplast transformation, VmNRPS12-knockout mutants were generated. PCR and Southern blot hybridization were applied to verify the positive mutant. Then genetic complementation was performed by transforming mutant protoplast with VmNRPS12 original gene. Finally, vegetative growth, conidiation and pathogenicity were examined.[Results] qRT-PCR analyses showed that VmNRPS12 was upregulated during the early infection stages of V. mali, and expressed remarkably at 48 hours post inoculation (138.6-fold). Compared with the wild type 03-8, VmNRPS12-knockout mutant showed no obvious change in vegetative growth and conidiation on PDA medium. Notably, the VmNRPS12-knockout mutant exhibited significantly reduced virulence on apple twigs. Furthermore, complementation of VmNRPS12 restored the pathogenecity of the VmNRPS12-knockout mutant approximately to the wild type 03-8.[Conclusion] VmNRPS12 contributes positively to the pathogenicity of V. mali.

Key words:Valsa mali;non-ribosomal peptide synthetase;gene knockout;complementation