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腺病毒DNA的体外复制。

Adenovirus DNA replication in vitro.

作者信息

Challberg M D, Kelly T J

出版信息

Proc Natl Acad Sci U S A. 1979 Feb;76(2):655-9. doi: 10.1073/pnas.76.2.655.

DOI:10.1073/pnas.76.2.655
PMID:284391
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC383008/
Abstract

A soluble extract from the nuclei of HeLa cells infected with adenovirus 5 (Ad5) carries out the semiconservative replication of exogenously added Ad5 DNA in vitro. Maximal DNA synthesis is observed when DNA-protein complex, isolated from Ad5 virions, is added as template. DNA-protein complex from virions of the closely related virus, adenovirus 2, is also active. In contrast, very little in vitro DNA synthesis is observed when deproteinized Ad5 DNA or DNA from a heterologous source (bacteriophage T7) is added as template. The product of the in vitro reaction consists of long Ad5 DNA strands that are hydrogen-bonded, but not covalently linked, to the input DNA template. During the course of the in vitro reaction, branched molecules with structural features identical to in vivo replication intermediates are formed. These findings support the conclusion that replication in the in vitro system closely resembles adenovirus DNA replication in vivo. The system provides an assay that should be useful for the purification and subsequent characterization of viral and cellular proteins involved in DNA replication.

摘要

从感染腺病毒5（Ad5）的HeLa细胞核中提取的可溶性提取物可在体外对外源添加的Ad5 DNA进行半保留复制。当添加从Ad5病毒粒子中分离出的DNA-蛋白质复合物作为模板时，可观察到最大程度的DNA合成。来自密切相关病毒腺病毒2的病毒粒子的DNA-蛋白质复合物也具有活性。相比之下，当添加脱蛋白的Ad5 DNA或来自异源来源（噬菌体T7）的DNA作为模板时，在体外观察到的DNA合成很少。体外反应的产物由长的Ad5 DNA链组成，这些链通过氢键与输入DNA模板相连，但不是共价连接。在体外反应过程中，形成了具有与体内复制中间体相同结构特征的分支分子。这些发现支持了体外系统中的复制与体内腺病毒DNA复制非常相似的结论。该系统提供了一种检测方法，可用于纯化和随后鉴定参与DNA复制的病毒和细胞蛋白。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b3eb/383008/ad68e1730998/pnas00002-0116-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b3eb/383008/27ad28cdb891/pnas00002-0115-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b3eb/383008/ad68e1730998/pnas00002-0116-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b3eb/383008/27ad28cdb891/pnas00002-0115-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b3eb/383008/ad68e1730998/pnas00002-0116-a.jpg

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