新疆苹果皱果类病毒(AFCVd)的检测与序列分析  

Detection of Apple fruit crinkle viroid in Xinjiang Apple and Its Specific Fragment Sequence Analysis

作　　者：赵英[1] 牛建新[1] 

机构地区：[1]石河子大学农学院园艺系,石河子832003

出　　处：《农业生物技术学报》2009年第1期164-169,共6页Journal of Agricultural Biotechnology

基　　金：国家自然科学基金资助项目(No.30360066);国家科技攻关计划引导项目(No.2003BA546C);兵团科委项目(No.NKB02SDXNK01SW)资助

摘　　要：采集新疆焉耆地区的老果园中的苹果(Malus domestica Brkn)树枝条、叶片和韧皮部,提取总RNA,根据Gen Bank中的AB104558序列设计引物,利用RT-PCR技术扩增出苹果皱果类病毒(Apple fruit crinkle viroid,AFCVd)特异条带,通过回收、克隆和测序,结果表明,获得的10条序列,已在GenBank中登录(登录号:EU031507～EU031516)。利用带有AFCVd目的DNA的质粒为模板,成功地合成了地高辛标记的cDNA探针,该探针能很好地用于AFCVd的检测。并利用原位RT-PCR技术做进一步检测,证明苹果叶片中有苹果皱果类病毒存在,主要分布在叶肉细胞的细胞核中。Branches, leaves and phloem of apple （Malus domestica Brkn） collected from an old orchard in Yanji and Xinjiang were used for total RNA isolation and specific fragments of Apple fruit crinkle viroid （AFCVd） were amplified by RT-PCR using the primers disigned according to the sequence AB 104558 on GenBank. Ten sequences were obtained through recovering, cloning and se- quencing, and also registered in GenBank （accession No. EU031507-EU031516）. Then cDNA probe laballed by Digoxigenin using plasmids containing targent DNA of AFCVd was composed and used for AFCVd detection. At the same time, in situ RT-PCR of AFCVd for further detection proved that AFCVd existed in apple leaf and mainly located in nucleus of mesophyll cell.

关 键 词：苹果皱果类病毒 原位RT-PCR 全序列分析 

分 类 号：S188[农业科学—农业基础科学]