苹果褪绿叶斑病毒的分子杂交检测  

Detection of Apple chorotic leaf spot virus by Dot- blot Hybridization

作　　者：杨洪一[1] 李昱莹 

机构地区：[1]东北林业大学生命科学学院,黑龙江哈尔滨150040

出　　处：《安徽农业科学》2013年第31期12324-12325,共2页Journal of Anhui Agricultural Sciences

摘　　要：[目的]探索检测苹果褪绿叶斑病毒的技术体系。[方法]利用CTAB法提取感病苹果叶片总RNA,以总RNA为模板,利用特异性引物,建立利用cDNA探针检测苹果褪绿叶斑病毒的技术体系。[结果]扩增出了预期片段,经克隆、测序证明该片段为苹果褪绿叶斑病毒的特异性片段。以带有特异片段的质粒为模板,利用PCR技术制备出地高辛标记的cDNA探针。[结论]以质粒为模板可高效制备大量探针。地高辛标记探针为非放射性标记探针,安全性较好,灵敏度较高,具有重要的应用价值。[ Objective ] The aim was to develop a system of detection for Apple chorotic leaf spot virus （ACILSV） with molecular hybridiza- tion. [ Method ] Total RNA was extracted from the leaves of infected apple by CTAB method, and the total RNA was used as the template to amplify the genome of ACLSV with the ACLSV specific primers. The system of detection of ACI.SV was developed by eDNA probe. [ Result] The predicted fragment was obtained, and the specific fragment was verified by cloning and sequencing. The digoxigenin-labeled eDNA probe is prepared by PCR using the plasmid with the specific fragment as a template. [ Conclusion ] The plasmid DNA was used as the template, then a number of probe could be effectively prepared. In addition, the digoxigenin-labeled probe was a non-radioactive one, so it was safe and was also high sensitive. The probe has an important significance for application.

关 键 词：苹果褪绿叶斑病毒 CDNA探针 检测 

分 类 号：S436.611.19[农业科学—农业昆虫与害虫防治]