Laboratory of Biotechnology, Green Chemistry Research Division, Research Institute of Green Science and Technology, Shizuoka University, 836 Ohya Suruga-ku, Shizuoka, 422-8529, Japan.
Laboratory of Biotechnology, Department of Applied Biological Chemistry, Faculty of Agriculture, Shizuoka University, 836 Ohya, Suruga-ku, Shizuoka, 422-8529, Japan.
Sci Rep. 2016 Aug 26;6:32283. doi: 10.1038/srep32283.
Autographa californica multiple nucleopolyhedrovirus (AcMNPV), an alphabaculovirus, has been widely utilized for protein expression in not only insect cells but also mammalian cells. AcMNPV is closely related to Bombyx mori nucleopolyhedrovirus (BmNPV), and nucleotide sequences of AcMNPV genes have high similarity with those of BmNPV. However, the transduction of BmNPV into mammalian cells has not been reported. In this study, we constructed a recombinant BmNPV (BmNPVΔbgp/AcGP64/EGFP) whose surface 64 kDa glycoprotein (BmGP64) was substituted with that from AcMNPV (AcGP64). BmNPVΔbgp/AcGP64/EGFP also carried an EGFP gene under the control of the CMV promoter. BmNPVΔbgp/AcGP64/EGFP successfully transduced HEK293T cells. In comparison, a control construct (BmNPVΔbgp/BmGP64/EGFP) which possessed BmGP64 instead of AcGP64 did not express EGFP in HEK293T cells. The transduction efficiency of BmNPVΔbgp/AcGP64/EGFP was lower than that of an AcMNPV based-BacMam GFP transduction control. This result indicates that AcGP64 facilitates BmNPV transduction into HEK293T cells. BmNPV can be prepared easily on a large scale because BmNPV can infect silkworm larvae without any special equipment, even though specific diet is needed for silkworm rearing. BmNPV gene transduction into mammalian cells can potentially be applied easily for gene delivery into mammalian cells.
苜蓿银纹夜蛾多核型多角体病毒(AcMNPV)是一种α杆状病毒,不仅在昆虫细胞中,而且在哺乳动物细胞中都广泛用于蛋白质表达。AcMNPV 与家蚕核型多角体病毒(BmNPV)密切相关,AcMNPV 基因的核苷酸序列与 BmNPV 的高度相似。然而,尚未报道 BmNPV 向哺乳动物细胞的转导。在这项研究中,我们构建了一种重组 BmNPV(BmNPVΔbgp/AcGP64/EGFP),其表面 64kDa 糖蛋白(BmGP64)被 AcMNPV 的糖蛋白(AcGP64)取代。BmNPVΔbgp/AcGP64/EGFP 还携带一个在 CMV 启动子控制下的 EGFP 基因。BmNPVΔbgp/AcGP64/EGFP 成功地转导了 HEK293T 细胞。相比之下,一个携带 BmGP64 而不是 AcGP64 的对照构建体(BmNPVΔbgp/BmGP64/EGFP)在 HEK293T 细胞中没有表达 EGFP。BmNPVΔbgp/AcGP64/EGFP 的转导效率低于基于 AcMNPV 的 BacMam GFP 转导对照。这一结果表明,AcGP64 促进了 BmNPV 向 HEK293T 细胞的转导。由于 BmNPV 可以感染家蚕幼虫而无需任何特殊设备,即使家蚕饲养需要特殊的饮食,因此可以很容易地大规模制备 BmNPV。BmNPV 基因向哺乳动物细胞的转导可能很容易应用于将基因递送到哺乳动物细胞中。