Department of Microbiology and Microbial Engineering, School of Life Sciences, Fudan University, Shanghai, China.
J Gene Med. 2013 Aug-Sep;15(8-9):282-90. doi: 10.1002/jgm.2722.
The baculovirus Autographa californica multiple nucleopolyhedrovirus has been widely explored as a transgene expression vector. Further improvement of the expression of the transgene is important for its application.
Bovine papillomavirus type 1 (BPV-1) cis-element upstream regulatory region (URR) and trans-elements E1, E2, were inserted into the baculovirus genome. The expression of reporter gene, enhanced green fluorescent protein gene (EGFP), and the persistence of viral genome was compared in several mammalian cell lines after virus transduction. The cytotoxicity of the recombinant viruses was also evaluated.
The recombinant baculovirus containing URR and E1, E2 genes showed significantly increased expression of EGFP in all cell lines tested, including HEK293, HeLa, BHK-21, CNE, CHO and MDCK cells. In HEK293 cells, the total production of EGFP was approximately five-fold higher than the control. The genome of virus with BPV-1 elements also persisted better than the control virus during the first few days post transduction. No obvious cytotoxicity was observed.
The coexistence of BPV-1 URR and E1, E2 was essential and sufficient to improve the performance of baculovirus with respect to mediating gene expression in various mammalian cells without major cytotoxicity. The results obtained in the present study facilitate the application of baculovirus as an efficient transgene vehicle for protein production and gene delivery.
杆状病毒 Autographa californica 多角体病毒已被广泛探索作为转基因表达载体。进一步提高转基因的表达对于其应用非常重要。
将牛乳头瘤病毒 1 型（BPV-1）顺式元件上游调控区（URR）和反式元件 E1、E2 插入杆状病毒基因组。比较了报告基因增强型绿色荧光蛋白基因（EGFP）在几种哺乳动物细胞系中转导病毒后的表达以及病毒基因组的持续存在。还评估了重组病毒的细胞毒性。
含有 URR 和 E1、E2 基因的重组杆状病毒在所有测试的细胞系中均显示出 EGFP 的表达显著增加，包括 HEK293、HeLa、BHK-21、CNE、CHO 和 MDCK 细胞。在 HEK293 细胞中，EGFP 的总产量比对照高约五倍。在转导后的头几天，含有 BPV-1 元件的病毒基因组的持续存在也优于对照病毒。未观察到明显的细胞毒性。
BPV-1 URR 和 E1、E2 的共存对于提高杆状病毒在各种哺乳动物细胞中介导基因表达的性能是必要且充分的，而不会产生明显的细胞毒性。本研究的结果为杆状病毒作为蛋白质生产和基因传递的有效转基因载体的应用提供了便利。
