应用RT-PCR和斑点杂交法检测新疆梨树上的苹果锈果类病毒  

Detection of apple scar skin viroid in pear cultivars by RT-PCR and spot hybridization in Xinjiang area

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,新疆石河子832003

出　　处：《果树学报》2007年第6期761-764,共4页Journal of Fruit Science

基　　金：国家自然科学基金(30360066);国家科技攻关计划引导项目(2003BA546C);兵团科委项目(NKB02SDXNK01SW)

摘　　要：利用自行设计的方法分离提取总RNA,通过一对特异引物,分别扩增出不同梨树品种上苹果锈果类病毒的特异片段,通过对库尔勒香梨品种上获得的特异片段回收、克隆和测序。结果表明,该片段长250bp,与已发表的AF421195同源性为99%,在此基础上利用克隆的ASSVd质粒,通过RT-PCR合成了生物素标记的cDNA探针,利用该探针对梨样品进行了斑点杂交检测,取得了很好的检测效果。进一步证明该反应体系能很好地用于梨树苹果锈果类病毒的RT-PCR检测。Total RNA was isolated using the method designed by ourselves. Full-length fragments of Apple scar skin viroid （ASSVd） have been amplified by one specific pair of primer from different pear samples. The ASSVd -specific fragment isolated from Korla＇s Xiangli pear cultivar has been recovered, cloned and sequenced, and the result showed its length was 250 bp representing the homology of 99% compared with AF421195 published on NCBI database. Then biotin-labelled cDNA probe was synthesized using cloned plasmids by RT-PCR. The probe has been used well for spot hybridization detection of ASSVd, which further proved RT-PCR can be successfully used for the detection of ASSVd in pear.

关 键 词：梨树 新疆 苹果锈果类病毒 RT—PCR CDNA探针 测序 

分 类 号：S661.2[农业科学—果树学]