摘要
采集新疆焉耆地区的老果园的苹果树枝条、叶片和韧皮部,利用自行设计的方法提取总RNA,根据GenBank中的X99487序列设计引物,利用RT-PCR技术扩增出苹果凹果类病毒(ADFVd)特异条带,通过回收克隆测序,结果表明,获得的7条序列,有5条同源性很高,均在98%以上,另外2条的同源性只有85%左右,这7条序列均已在GenBank中登录(登录号:EU031497_EU031503)。在此基础上利用插入ADFVd目的DNA的质粒为模板,成功合成了地高辛标记的cDNA探针,该探针能很好地用于ADFVd的检测。同时利用原位RT-PCR技术,做了进一步检测,证明苹果叶片中有苹果凹果类病毒存在,主要分布在叶肉细胞的细胞核中。
The apple branches, leaves and phloem collected from an old orchard in Yanji, Xinjiang were used as materials for total RNA isolation using the method designed. Specific fragments of apple dimple fruit viroid （ADFVd）were amplified by RT-PCR using the primers designed according to the sequence X99489 on GenBank. 7 sequences were obtained through recovering and sequencing; 5 sequences had high homology over 98%,the other two sequences only- about 85%, also registered in GenBank （acc. No. EU031497 EU031503 ）. The cDNA probe labelled by digoxigenin was prepared from plasmids containing targent DNA of ADFVd and it can be successfully- used for ADFVd detection. At the same time, the detection results by in situ RT-PCR proved the presence of ADFVd in apple leaf, and it mainly located in nucleus of mesophyll cell.
出处
《果树学报》
CAS
CSCD
北大核心
2008年第5期682-685,F0004,共5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)
