Overexpression of Cordyceps militaris Cmcns1/cns2 gene to produce cordycepin results in increased resistance to Ralstonia solawacearum, Alternaria alternate and TMV in tobacco(Nicotiana tabacum L.).

Cordycepin (COR), the first nucleoside antibiotic isolated from fungi, exhibits antibacterial, antitumor, antiviral, and anti-inflammatory activities. In this study, we constructed a Cmcns1/cns2 fusion gene vector and transformed tobacco using Agrobacterium tumefaciens-mediated genetic transformation. The results showed that cordycepin was detected in transgenic tobacco plants, with Cmcns1/cns2 expression confirmed and the highest expression level observed in line OE1. This suggests a direct correlation between COR production and Cmcns1/cns2 gene expression levels. Importantly, following infection with viral or bacterial pathogens, the Cmcns1/cns2-overexpressing tobacco plants exhibited fewer wilting lesions compared to wild-type plants, indicating that Cmcns1/cns2 expression contributes to COR production, thereby enhancing disease resistance. We measured malondialdehyde (MDA) content, as well as superoxide dismutase (SOD) and peroxidase (POD) activity, before and after treatment. The results demonstrated that MDA content in transgenic tobacco was lower than pre-treatment levels, whereas SOD and POD activities were higher than pre-treatment values. Furthermore, expression levels of disease resistance genes NtADR1, NtNPR1, and NtNRG1 in transgenic line OE1 were significantly elevated compared to wild-type plants following treatment with Ralstonia solanacearum and Alternaria alternata. This confirms that the transgenic Cmcns1/cns2 gene enhances disease resistance in tobacco, presenting a new approach for breeding plants with improved disease resistance.