Ryan K W, Mackow E R, Chanock R M, Lai C J
Virology. 1986 Oct 15;154(1):144-54. doi: 10.1016/0042-6822(86)90437-x.
Simian cells permissive for influenza A virus infection were stably transformed with a full-length cloned influenza A nucleoprotein gene under the control of an inducible metallothionein promoter and linked to a dihydrofolate reductase gene to facilitate cell selection. Transformed cells synthesized a virus-specific nucleoprotein which was indistinguishable from the nucleoprotein synthesized in virus-infected cells with respect to molecular weight and intracellular localization. It was estimated that transformed cells produced only 1% of the amount of nucleoprotein synthesized in simian cells infected with influenza A virus. Nonetheless, when transformed cells were infected with influenza virus mutants which synthesized temperature-sensitive nucleoprotein, protein expressed by the cloned gene was able to complement the synthesis of plus-strand and minus-strand viral RNA for one mutant and only plus-strand synthesis for another mutant. This indicated that the influenza A nucleoprotein expressed in the transformed cells exhibited functional activity.
对甲型流感病毒感染敏感的猿猴细胞，在可诱导的金属硫蛋白启动子的控制下，用全长克隆的甲型流感病毒核蛋白基因进行稳定转化，并与二氢叶酸还原酶基因相连以促进细胞筛选。转化细胞合成了一种病毒特异性核蛋白，其分子量和细胞内定位与病毒感染细胞中合成的核蛋白没有区别。据估计，转化细胞产生的核蛋白量仅为感染甲型流感病毒的猿猴细胞中合成量的1%。尽管如此，当用合成温度敏感核蛋白的流感病毒突变体感染转化细胞时，克隆基因表达的蛋白能够为一个突变体互补正链和负链病毒RNA的合成，为另一个突变体仅互补正链合成。这表明在转化细胞中表达的甲型流感病毒核蛋白具有功能活性。
