新疆苹果皱果类病毒(AFCVd)的检测与序列分析

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GenBank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。

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

Branches, leaves and phloem of apple (Malus domestica Brkn) collected from an old orchard in Yanji, Xinjiang were used for total RNA isolation specific fragments of Apple fruit crinkle viroid (AFCVd) were amplified by RT-PCR using the primers disigned according to the sequence AB104558 on GenBank. Ten sequences were obtained through recovering, cloning and sequencing, 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 the presence of AFCVd in apple leaf, mainly located in nucleus of mesophyll cell.