Objective: We developed an efficient method of gene knockout in Verticillium dahliae, an important soil-borne fungal pathogen that causes cotton vascular wilt diseases.
Methods: By using fusion PCR, we constructed gene knockout vectors. By using Agrobacterium tumefaciens-mediated transformation and applying a herpes simplex virus thymidine kinase (HSVtk) gene in T-DNA as a conditional lethal gene to counter-select against ectopic transformants, we developed an efficient method to select gene knockout transformants.
Results: Gene knockout frequency for ADE4 and ChsV was 87% and 44% , respectively.
Conclusion: We developed an efficient tool for gene knockout in Verticillium dahliae, which would help clarify the infection mechanism of this fungal pathogen.