CTV强弱毒分离株中p20的表达特征及其对SA通路相关基因的影响  

Expression Characteristics p20 of the Severe and Mild Strains of CTV and Its Effect on SA Pathway Related Genes

作　　者：陈琪 颜复林 姜俊遥 王洪苏 陈静 刘露勤 梁佳梅 刘金香[1] CHEN Qi;YAN Fulin;JIANG Junyao;WANG Hongsu;CHEN Jing;LIU Luqin;LIANG Jiamei;LIU Jinxiang(Citrus Research Institute,Southwest University,Chongqing 400712,China;Sichuan Academy of Chinese Medicine Sciences,Chengdu 610041,China)

机构地区：[1]西南大学柑桔研究所,重庆400712 [2]四川省中医药科学院,成都610041

出　　处：《西南大学学报（自然科学版）》2024年第9期45-54,共10页Journal of Southwest University(Natural Science Edition)

基　　金：国家自然科学基金项目(31972237);国家重点研发计划项目(2021YFD1400800)。

摘　　要：为明确柑橘衰退病毒(CTV)强弱毒分离株中沉默抑制子p20的表达特性及其对SA通路关键基因的转录水平,采用RT-PCR、 cDNA克隆及基因组测序等技术获得p20基因序列,运用MEGA7和DNAMAN软件对CTV强弱毒分离株的p20氨基酸序列进行系统进化树构建和比对分析.结果表明,中国弱毒分离株CTLJ与T30,T358之间的亲缘关系最近,同源性为98.35%.进一步通过Western blot分析表明,CTV强分离株CT14A在甜橙嫩皮中p20的表达水平显著高于弱毒分离株CTLJ,然后探究了这两种强弱毒株间p20的沉默抑制能力是否有差异,结果表明强毒分离株CT14A和弱毒分离株CTLJ的p20蛋白均具有沉默抑制子活性,但是二者沉默抑制活性无显著差异.最后,RT-qPCR技术定量分析结果表明,烟草中瞬时表达p20会诱导SA信号通路关键基因PR1,PR1a和NPR1的转录水平发生变化.To elucidate the expression characteristics of silencing suppressor p20 of severe and mild isolates of citrus tristeza virus(CTV),and its effect on the transcription level of key genes in SA pathway,the techniques of RT-PCR,cDNA cloning and gemomic sequencing were used to obtain p20 gene sequence.The p20 amino acid sequences of CTV severe and mild isolates were analyzed by software DNAMAN and MEGA7.The results showed that the mild strain CTLJ had the closest genetic relationship with T30 and T358,with a homology of 98.35%.Furthermore,Western blot analysis showed that the expression level of p20 protein of CT14A of severe CTV isolate in tender bark of sweet orange was significantly higher than that of CTLJ of mild CTV isolate.The difference on silencing inhibition ability of p20 between the two isolates was explored.The results showed that both severe isolate CT14A and mild isolate CTLJ had silencing inhibitor activity,but there was no significant difference between them.Finally,plant transient expression vector was constructed and transformed to Nicotiana benthamiana by Agrobacterium tumefaciens The RT-qPCR was used to analyze the transcriptional levels of key genes in SA pathway.The result showed that transient expression of p20 in N.benthamiana caused changes in the transcriptional levels of PR1,PR1a and NPR1.

关 键 词：柑橘衰退病毒 p20蛋白 沉默抑制子 水杨酸 

分 类 号：S436.66[农业科学—农业昆虫与害虫防治]