日本三株斜纹夜蛾核型多角体病毒的增殖特性及其多角体蛋白基因的序列分析

利用斜纹夜蛾Spodoptera litura培养细胞，对近年来在日本本州、九州和四国等地发现并筛选出的对斜纹夜蛾幼虫具有强烈杀虫活性的3株斜纹夜蛾核型多角体病毒(SpltMNPV)（K-3、G1-2和G10-3）进行了生物学活性和分子生物学的初步研究,克隆了多角体蛋白基因,并进行了序列分析和比较。结果表明:（1）SpltMNPV日本分离株K-3、G1-2和G10-3分别具有不同的特征性酶切图谱，分别属于3种基因型（A型、B型和C型）； （2）3个分离株的芽生型病毒（budded virus）产生能力和多角体产生能力有差异，免疫印迹分析表明，多角体蛋白的分子量也不同；（3）日本株SpltMNPV核型多角体蛋白结构基因由747个核苷酸编码序列（编码249个氨基酸）组成，其序列与中国株SpltMNPV的同源性为98.9%,与其他6种核型多角体病毒有较高的同源性（61.7%～74.2%）,但其5′端侧翼序列(nt-1～-100)与AcMNPV和BmNPV相比差异显著,在对该基因表达调控起决定性作用的8个高度保守核苷酸序列中（nt-44～-51）有2处发生自然突变。

Proliferation characteristics of three Japanese strains of Spodoptera litura multicapsid nucleopolyhedrovirus in cultural cells and sequence analysis of their polyhedrin genes

Biological and molecular characteristics of three strains (K-3, G1-2 and G-10) of Spodoptera litura multicapsid nucleopolyhedrovirus (SpltMNPV), isolated from Honshu, Kyushu and Shikoku, Japan, which had strong pesticide activity to S. litura, were primarily studied using the cultured cell strain of S. litura. The polyhedrin genes of the three strains was cloned and sequenced. The results showed that: (1) the three Japanese strains K-3, G1-2 and G10-3 had different restriction fragmentation patterns and belonged to the nucleic acid type A, B and C of SpltMNPV respectively. (2) There are differences in the outputs of budded virus (BV) and polyhedron between the three strains. According to the analysis of immunological blotting, the molecular weights of polyhedrin in the three strains were different. (3) The polyhedrin gene of the Japanese SpltMNPV is composed of 747 nucleic acids, which codes a peptide of 249 amino acids. The nucleic acid sequences homology is as high as 98.9% compared with the Chinese strain, and 61.7%-74.2% with other six NPVs compared. However, in the 5' upstream sequence (nt-1-100), SpltMNPV differs significantly with AcMNPV and BmNPV. There are 2 point mutations in the sequence with 8 highly conservative nucleic acids (nt-44-51), which is the domain of the promoter. Interestingly, in spite of only one mutation occurred in the polyhedron open reading frame of SpltMNPV C type, the western blot result showed that the molecular weight of polyhedron of SpltMNPV C type is apparently different from that of A type and B type.