五味子叶枯病菌细极链格孢GFP荧光标记及侵染过程OA

Leaf blight of Schisandra chinensis,caused by Alternaria tenuissima,is an important disease in commercial fields.To observe the infection process of the pathogen,a GFP fluorescent labeling vector pBS-HYG-GFP was constructed using homologous recombination technology and the protoplast preparation conditions were optimized(hyphae aged 36 h enzymolysed with 0.01 g/mL driselase,0.01 g/mL lyticase and 0.01 g/mL snailase at 28℃for 2.5 h).The protoplast yield reached 1.14×106/mL.Following genetic transformation,a stable fluorescent strain SccA071GFP was obtained that exhibited no significant differences from the wild-type strain in terms of growth rate,spore production capacity and pathogenicity.Infection process observation revealed that conidia attached and germinated on the leaf surface,with germ tubes developing into primary hyphae.The hyphae preferentially extended along the depressions on the leaf surface and mainly invaded the mesophyll tissue through stomata or intercellular spaces in 36 hours after inoculation.In 96 hours after inoculation,the hyphae spread in a network within the mesophyll tissue,and extensively branched and densely colonized the tissue,completing the entire process of penetration,incubation,and symptom development in 120 hours after inoculation.In this study,an efficient and stable genetic transformation system was first established for the pathogen causing leaf blight in S.chinensis.The findings provide a stable and efficient genetic transformation system for identifying functional genes in the pathogen,and advantage in-depth investigations into pathogen infection and pathogenesis mechanisms.