Replication of plant RNA virus genomes in a cell-free extract of evacuolated plant protoplasts
Replication of plant RNA virus genomes in a cell-free extract of evacuolated plant protoplasts
复制标题
DOI：
10.1073/pnas.0307131101
复制
发表时间：
2004-02-17
影响因子：
11.1
通讯作者：
Ishikawa, M
中科院分区：
文献类型：
--
作者：
Komoda, K;Naito, S;Ishikawa, M
真核正链RNA病毒基因组的复制是一个复杂的过程，涉及多种病毒和宿主蛋白以及胞内膜。在这里，我们报告了一个无细胞系统，在体外复制这一过程。该系统使用含有膜的未感染植物原生质体提取物，其中液泡已通过Percoll梯度离心去除。我们证明该系统支持番茄花叶病毒和其他两种植物正链RNA病毒的翻译、负链RNA合成、基因组RNA复制和亚基因组RNA转录。RNA合成依赖于基因组RNA的翻译，产生与病毒相关的RNA物种，类似于在体内产生的RNA物种。该系统将有助于阐明这些病毒的基因组复制机制。
The replication of eukaryotic positive-strand RNA virus genomes occurs through a complex process involving multiple viral and host proteins and intracellular membranes. Here we report a cell-free system that reproduces this process in vitro. This system uses a membrane-containing extract of uninfected plant protoplasts from which the vacuoles had been removed by Percoll gradient centrifugation. We demonstrate that the system supported translation, negative-strand RNA synthesis, genomic RNA replication, and subgenomic RNA transcription of tomato mosaic virus and two other plant positive-strand RNA viruses. The RNA synthesis, which depended on translation of the genomic RNA, produced virus-related RNA species similar to those that are generated in vivo. This system will aid in the elucidation of the mechanisms of genome replication in these viruses.