通过Hirt提取和Southern印迹分析腺相关病毒的复制中间体
Analysis of Replicative Intermediates of Adeno-associated Virus through Hirt Extraction and Southern Blotting.
作者信息
Bardelli Martino, Zarate-Perez Francisco, Agundez Leticia, Jolinon Nelly, Michael Linden R, Escalante Carlos R, Henckaerts Els
机构信息
Department of Infectious Diseases, King's College London, London, United Kingdom.
Department of Physiology and Biophysics, Virginia Commonwealth University School of Medicine, Richmond VA, USA.
出版信息
Bio Protoc. 2017 May 5;7(9). doi: 10.21769/BioProtoc.2271.
Abstract
Adeno-associated virus (AAV) is a small single-stranded DNA virus that requires the presence of a helper virus, such as adenovirus or herpes virus, to efficiently replicate its genome. AAV DNA is replicated by a rolling-hairpin mechanism (Ward, 2006), and during replication several DNA intermediates can be detected. This detailed protocol describes how to analyze the AAV DNA intermediates formed during AAV replication using a modified Hirt extract (Hirt, 1967) procedure and Southern blotting (Southern, 1975).
摘要
腺相关病毒（AAV）是一种小型单链DNA病毒，它需要辅助病毒（如腺病毒或疱疹病毒）的存在才能有效地复制其基因组。AAV DNA通过滚发夹机制进行复制（沃德，2006年），并且在复制过程中可以检测到几种DNA中间体。本详细方案描述了如何使用改良的Hirt提取物（Hirt，1967年）方法和Southern印迹法（Southern，1975年）分析AAV复制过程中形成的AAV DNA中间体。
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