van Oers M M, Malarme D, Jore J M, Vlak J M
Department of Virology, Wageningen Agricultural University, The Netherlands.
Arch Virol. 1992;123(1-2):1-11. doi: 10.1007/BF01317134.
Two major late proteins, polyhedrin and p10, are synthesized in large quantities in baculovirus infected insect cells. This and the fact that both proteins are dispensable for virus replication, form the basis for the use of these viruses as vector for foreign gene expression. To address the question whether the Autographa californica nuclear polyhedrosis virus p10 promotor-driven expression is influenced by the concurrent expression of the polyhedrin gene, several recombinants were constructed with various deletions in the polyhedrin gene. The Escherichia coli lacZ gene was used as a marker to allow direct comparison between p10 and polyhedrin-driven expression. None of the deletions in the polyhedrin gene did result in higher expression of the p10 promoter-controlled gene. This suggested that the transcriptional and/or translational activity of the p10 and polyhedrin gene are independently regulated. To compare the level of polyhedrin and p10 promoter driven expression, recombinants with the lacZ gene cloned behind either promoter were studied. No significant difference in level of expression was observed. In cells infected with a recombinant with the lacZ gene present behind both promoters a reduced level of expression was observed, whereas a considerable increase was expected. This may be due to instability of the viral genome, as two copies of the lacZ gene were present.
两种主要的晚期蛋白,多角体蛋白和p10,在杆状病毒感染的昆虫细胞中大量合成。这一点以及这两种蛋白对于病毒复制都是非必需的这一事实,构成了将这些病毒用作外源基因表达载体的基础。为了探讨苜蓿银纹夜蛾核型多角体病毒p10启动子驱动的表达是否受多角体蛋白基因同时表达的影响,构建了几种在多角体蛋白基因中有不同缺失的重组体。大肠杆菌lacZ基因用作标记,以便直接比较p10和多角体蛋白驱动的表达。多角体蛋白基因中的任何缺失均未导致p10启动子控制基因的更高表达。这表明p10和多角体蛋白基因的转录和/或翻译活性是独立调节的。为了比较多角体蛋白和p10启动子驱动的表达水平,研究了将lacZ基因克隆在任一启动子后面的重组体。未观察到表达水平的显著差异。在用两个启动子后面都带有lacZ基因的重组体感染的细胞中,观察到表达水平降低,而预期会有相当大的增加。这可能是由于病毒基因组的不稳定性,因为存在两个lacZ基因拷贝。