Reovirus RNA is infectious.
Reovirus RNA is infectious.
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呼肠孤病毒RNA具有传染性。
DOI：
10.1016/0042-6822(90)90153-i
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发表时间：
1990
期刊：
影响因子：
3.7
通讯作者：
Joklik,WK
中科院分区：
文献类型：
--
作者：
Roner,MR;Sutphin,LA;Joklik,WK
呼肠孤病毒RNA具有感染性的条件已经确定。简而言之，将呼肠孤病毒血清型3（ST 3病毒）的单链（正链，ss）和/或双链（ds）RNA与其中已翻译ss或解链dsRNA的兔网织红细胞裂解物一起脂质转染到L929小鼠成纤维细胞中。8小时后，用辅助病毒ST 2呼肠孤病毒感染细胞。24或48小时后收获病毒产量。在这些条件下，产生5天形成空斑的病毒，所有这些病毒都是ST 3病毒; ST 2病毒仅在12天后形成空斑。后代中不存在抑制物。病毒产量约为0.2 PFU/细胞;免疫荧光测定显示，该子代来自约4%的细胞。双链RNA的感染性是ssRNA的20倍; ds和ssRNA一起产生的感染性病毒是单独dsRNA的10倍，原因是dsRNA大大增加了ssRNA的感染性。所有种类的ss和dsRNA都需要这种加和效应的运作。致敏的兔网织红细胞裂解物不是必需的，但可使病毒产量增加100倍。它的活性与翻译进行的时间成正比;然而，这种活性不仅仅是由于新合成的蛋白质，因为翻译后RNA的破坏会消除活性，而这种活性不能通过简单地添加更多的RNA来恢复。所有种类的RNA的翻译都是必不可少的。尽管当不同基因型的ss和dsRNA一起脂质转染时不形成阻遏物，但不同基因型的dsRNA的混合物确实产生阻遏物。对于ssRNA的这种混合物也是如此。这些发现将允许将新的或改变的基因组片段引入呼肠孤病毒基因组中。它们为呼肠孤病毒基因组片段上的标记和分类信号的鉴定、呼肠孤病毒蛋白的功能结构域的表征以及呼肠孤病毒作为表达载体的开发开辟了道路。
Conditions under which reovirus RNA is infectious have been worked out. In brief, single-stranded (plus-stranded, ss) and/or double-stranded (ds) RNA of reovirus serotype 3 (ST3 virus) is lipofected into L929 mouse fibroblasts together with a rabbit reticulocyte lysate in which ss or melted dsRNA has been translated. After 8 hr the cells are then infected with a helper virus, ST2 reovirus. Virus yields are harvested 24 or 48 hr later. Under these conditions virus that forms plaques by 5 days is produced, all of which is ST3 virus; ST2 virus forms plaques only after 12 days. No reassortants are present among the progeny. The virus yields are about 0.2 PFU/cell; immunofluorescence assays show that this progeny is derived from about 4% of the cells. Double-stranded RNA is 20 times as infectious as ssRNA; ds and ssRNA together yield 10 times as much infectious virus as dsRNA alone, the reason being that dsRNA greatly increases the infectiousness of ssRNA. All species of both ss and dsRNA are required for the operation of this additive effect. The primed rabbit reticulocyte lysate is not essential, but increases virus yields by 100-fold. Its activity is proportional to the time for which translation has proceeded; however, this activity is not due solely to newly synthesized proteins because destruction of the RNA following translation abolishes activity which cannot be restored by simple addition of more RNA. Translation of all species of RNA is essential. Whereas no reassortants are formed when ss and dsRNA of different genotypes are lipofected together, mixtures of dsRNAs of different genotypes do yield reassortants. The same is true for such mixtures of ssRNA. These findings will permit the introduction of new or altered genome segments into the reovirus genome. They open the way to the identification of encapsidation and assortment signals on reovirus genome segments, the characterization of functional domains on reovirus proteins, and the development of reovirus as an expression vector.