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羧基转运蛋白基因是近年来分离出的新基因,在虫生真菌中可能与穿透昆虫体壁时的能量代谢有关。构建了针对该基因的双链RNA干扰载体,采用芽生孢子转化法将载体质粒转入球孢白僵菌,并通过RT-PCR检测转化前后BbJEN1的表达情况。RT-PCR检测结果显示,球孢白僵菌转化子BbJEN1基因的表达水平显著下降,表明干扰载体具有明显的沉默效应。在两种培养基上,转化子与原始菌株的生长速度和产孢量差异均不显著,而分生孢子萌发则显著滞后于原始菌株。使用马尾松毛虫幼虫对转化子和原始菌株进行生物测定,原始菌株的半致死浓度、半致死剂量和半致死时间分别为2.79×106个孢子/mL、84.12个孢子/mm2和6.49d,而转化子的半致死浓度、半致死剂量和半致死时间则分别增加到1.27×107个孢子/mL、382.92个孢子/mm2和8.09d,毒力显著下降。由此表明BbJEN1基因与球孢白僵菌分生孢子发芽以及毒力均有关。
Carboxyltransferase gene is a new gene isolated in recent years, which may be related to the energy metabolism when penetrating insect body wall in entomogenous fungi. The double-stranded RNA interference vector was constructed. The vector plasmid was transformed into Beauveria bassiana by using sporulation method. The expression of BbJEN1 was detected by RT-PCR. The results of RT-PCR showed that the expression of BbJEN1 gene of Beauveria bassiana decreased significantly, which indicated that the interference vector had a significant silencing effect. On both media, the growth rate and sporulation of transformants were not significantly different from those of the original strain, while conidial germination lags behind that of the original strain significantly. The semi-lethal concentration, lethality and lethality of the original strain were 2.79 × 106 spores / mL, 84.12 spores / mm2 and 6.49 d, respectively, for the bioassay of the transformant and the original strain using Dendrolimus punctatus larvae, The lethal concentration, the lethal dose and the lethal time increased to 1.27 × 107 spores / mL, 382.92 spores / mm2 and 8.09d respectively, and the virulence decreased significantly. The results showed that the BbJEN1 gene was associated with the conidiospore germination and virulence of B. bassiana.