利用宿主范围扩大的重组杆状病毒高效表达外源基因。

High-level expression of a foreign gene by a recombinant baculovirus with an expanded host range.

机构信息

Division of Applied Biology and Chemistry, College of Agriculture and Life Sciences, Seoul National University, Suwon, 441-744, Korea.

出版信息

Cytotechnology. 2000 Feb;32(2):87-92. doi: 10.1023/A:1008166310368.

DOI:10.1023/A:1008166310368

PMID:19002970

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC3449685/

Abstract

The usefulness of host range expanded viruses as an expressionvector system was investigated by following the expression ofthe E. coli lacZ gene. The host range expanded recombinantviruses were obtained from Sf-21 or BmN-4 cells coinfected withAutographa californica and Bombyx mori nuclearpolyhedrosis viruses. Among the host range expanded viruses,RecB-8 and RecS-B6 have similar enzyme digestion profiles butdifferent infection characteristics in cells. Therefore, tostudy the foreign gene expression efficiency of these twoviruses, we constructed recombinant viruses RecB8-LacZ andRecSB6-LacZ containing the lacZ gene instead of the polyhedringene. Also, the host range expanded recombinant AcNPV, Bac-BH,containing lacZ gene in the polyhedrin gene locus was constructedby substitution of the 0.6 kb region within the helicase gene ofBacPAK6 with that of BmNPV. beta-Galactosidase expressionefficiency by these viruses were determined and compared in Sf-21and BmN-4 cells. The result showed that Bac-BH has highexpression efficiency only in Sf-21 cells, whereas RecB8-LacZhas high expression efficiency both in Sf-21 and BmN-4 cells.Also, in BmN-4 cells, beta-galactosidase expressionefficiency of RecB8-LacZ was higher than that of recombinantBmNPV (BmK1-LacZ containing lacZ gene in polyhedrin gene locus).In addition, the expression efficiency was not correlated withvirus titer.

摘要

研究了宿主范围扩展病毒作为表达载体系统的有用性，方法是跟踪大肠杆菌 lacZ 基因的表达。从 Sf-21 或 BmN-4 细胞中共同感染美洲棉铃象鼻虫和家蚕核多角体病毒获得了宿主范围扩展的重组病毒。在宿主范围扩展的病毒中，RecB-8 和 RecS-B6 具有相似的酶切图谱，但在细胞中的感染特性不同。因此，为了研究这两种病毒的外源基因表达效率，我们构建了含有 lacZ 基因而不是多角体基因的重组病毒 RecB8-LacZ 和 RecSB6-LacZ。此外，通过用 BmNPV 的螺旋酶基因内的 0.6 kb 区域取代 BacPAK6 中的螺旋酶基因，构建了含有 lacZ 基因的宿主范围扩展的重组 AcNPV Bac-BH。在 Sf-21 和 BmN-4 细胞中测定了这些病毒的β-半乳糖苷酶表达效率并进行了比较。结果表明，Bac-BH 仅在 Sf-21 细胞中具有高表达效率，而 RecB8-LacZ 在 Sf-21 和 BmN-4 细胞中均具有高表达效率。此外，在 BmN-4 细胞中，RecB8-LacZ 的β-半乳糖苷酶表达效率高于含有多角体基因座 lacZ 基因的重组 BmNPV（BmK1-LacZ）。此外，表达效率与病毒滴度无关。