M olecular Cloning and Sequence Analysis of Dendrolimus punctatus cytoplasmic polyhedrosis virus NS5 Protein Gene

WEN Li; ZHANG Jia—min; WANG Qiong; HU Jian-fang; HU Yuan-yang

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February 2003

WEN Li, ZHANG Jia—min, WANG Qiong, HU Jian-fang and HU Yuan-yang. M olecular Cloning and Sequence Analysis of Dendrolimus punctatus cytoplasmic polyhedrosis virus NS5 Protein Gene[J]. Virologica Sinica, 2003, 18(1): 49-53.

Citation: WEN Li, ZHANG Jia—min, WANG Qiong, HU Jian-fang, HU Yuan-yang. M olecular Cloning and Sequence Analysis of Dendrolimus punctatus cytoplasmic polyhedrosis virus NS5 Protein Gene .VIROLOGICA SINICA, 2003, 18(1) : 49-53.

马尾松毛虫质型多角体病毒NS5蛋白基因的 cDNA克隆及序列分析

1.
武汉大学病毒研究所，湖北武汉430072

摘要

通过对马尾松毛虫质型多角体病毒的增殖、纯化，获得一株单一类型的质型多角体病毒。提纯的病毒粒子经SDS一热酚法抽提。在使用低熔点琼脂糖凝胶电泳分离基因组dsRNA，回收纯化第九片段S9。SgRNA双链经高温变性，使用正反两种引物逆转录合成cDNA双链，使用同样的引物经PCR扩增后，克隆在pMDI8-T载体上。利用两种引物组合，获得了大小两个亚克隆片段。序列测定结果表明，较小亚克隆片段长405bp，较大亚克隆片段长677bp，经过序列拼接，得到一个977bp的序列，其中包含一个963bp大的开放阅读框(ORF)。推测 DpCPVS9基因编码一个320个氨基酸的多肽，分子质量35．66kDa。和家蚕1型质型多角体病毒的I株(BmCPV—I I strain)位于第九片段的NS5蛋白基因相比较，核苷酸和编码氨基酸序列同源性分别为86．0％和93．4％。
- 马尾松毛虫
- , 质型多角体病毒
- , 第九片段(S9)
- , NS5蛋白

M olecular Cloning and Sequence Analysis of Dendrolimus punctatus cytoplasmic polyhedrosis virus NS5 Protein Gene

1.
Institute of Virology。Wuhan University， uhan 430072，China

Abstract

For the synthesis of cDNA of Dendrolimus punctaus CPV (DpCPV)NS5 protein，the primers were designed on the basis of the RNA sequmce of BmCPV一1(I strain)segment 9．After the RT—PCR the amplified cDNA was cloned into the pMD18一T． NS5 protein gene of DpCPV was found to be 963 nucleotides in length with a single open reading frame in one strand capable of coding a predicted pro— tein of 320 residues(Mw of 35．66kDa)． Comparison of the nucleotide sequences of the NS5 gene of DpCPV with those of BmCPV—I(1 strain)showed that nucleotides homology iS 86．0％， but there are only 2 1 amino acid differences between the two CPVs．
- Dendrolimus punctatus
- , Cypovirus
- , Segment 9
- , NS5 protein

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通讯作者: 陈斌, bchen63@163.com

1.
沈阳化工大学材料科学与工程学院沈阳 110142

M olecular Cloning and Sequence Analysis of Dendrolimus punctatus cytoplasmic polyhedrosis virus NS5 Protein Gene

1. Institute of Virology。Wuhan University， uhan 430072，China

Abstract: For the synthesis of cDNA of Dendrolimus punctaus CPV (DpCPV)NS5 protein，the primers were designed on the basis of the RNA sequmce of BmCPV一1(I strain)segment 9．After the RT—PCR the amplified cDNA was cloned into the pMD18一T． NS5 protein gene of DpCPV was found to be 963 nucleotides in length with a single open reading frame in one strand capable of coding a predicted pro— tein of 320 residues(Mw of 35．66kDa)． Comparison of the nucleotide sequences of the NS5 gene of DpCPV with those of BmCPV—I(1 strain)showed that nucleotides homology iS 86．0％， but there are only 2 1 amino acid differences between the two CPVs．