Cloning，Sequencing and Expressing of S Segment Coding Gene of Seoai Virus R22 Strain

TAO Zhi-Hui; DONG Guan-Mu; SHU Yong-Xin

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August 1999

TAO Zhi-Hui, DONG Guan-Mu and SHU Yong-Xin. Cloning，Sequencing and Expressing of S Segment Coding Gene of Seoai Virus R22 Strain[J]. Virologica Sinica, 1999, 14(4): 322-327.

Citation: TAO Zhi-Hui, DONG Guan-Mu, SHU Yong-Xin. Cloning，Sequencing and Expressing of S Segment Coding Gene of Seoai Virus R22 Strain .VIROLOGICA SINICA, 1999, 14(4) : 322-327.

肾综合征出血热病毒R22株核蛋白基因的克隆序列分析及其表达

1.
中国药品生物制品检定所，北京100050

摘要

为研究肾综合征出血热病毒疫苗侯选株R22核蛋白的结构与特性，应用逆转录PCR扩增了R22编码区基因，并将扩增产物克隆于pET-3a表达质粒，测得R22株S片段编码区序列为1290个核苷酸。比较分析表明与Seoul型同源性高达96．2％，而与Hantaan型同源性仅为71．0％，与用血清学分型的结果一致。将克隆的pET－R22NP转化到BL21后，IPTG诱导得到较高效的表达，产物纯化后进行Westernblot分析，结果表达的NP仅与NP蛋白特异的单克隆抗体A35和3D9反应，而与G2蛋白特异的3D8不反应。表达的产物为汉坦病毒的诊断提供了特异性抗原。
- 肾综合征出血热病毒
- , 核蛋白
- , 基因克隆
- , 序列分析
- , 表达

Cloning，Sequencing and Expressing of S Segment Coding Gene of Seoai Virus R22 Strain

1.
National Institute for the Control Pharmaceutical and biological

Abstract

For studing the structure and characteristic of nucleoprotein (NP) of Seoul virus R22strain, which is the seed virus of HFRS candidate vaccine, the NP gene coding region of the R22S segment was amplificated by RT-PCR and then cloned it into pET-3a expression plasmid. Se-quence assay showed that the NP gene contains 1290 nucleotides, which has homology of 96. 2%with well known Seoul virus (SR-11 ) and 71. 0% with Hantaan virus (76-118). These resultscoincide with previous Serological testings. The cloned NP gene has been transformed into E. coliBL21 and it could give a high expression when induced by IPTG. Western blot and ELISA assayof the purified products showed that expressed NP only reacted with NP-specific MAbs A35 and3D9, but not with G2-specific MA 3D8. The expressed NP can be used as a specific antigen forclinical diagnosis.
- Seoul virus
- , Nucleoprotein(NP)
- , Cloning
- , Sequencing
- , Expression

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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

Cloning，Sequencing and Expressing of S Segment Coding Gene of Seoai Virus R22 Strain

1. National Institute for the Control Pharmaceutical and biological

Abstract: For studing the structure and characteristic of nucleoprotein (NP) of Seoul virus R22strain, which is the seed virus of HFRS candidate vaccine, the NP gene coding region of the R22S segment was amplificated by RT-PCR and then cloned it into pET-3a expression plasmid. Se-quence assay showed that the NP gene contains 1290 nucleotides, which has homology of 96. 2%with well known Seoul virus (SR-11 ) and 71. 0% with Hantaan virus (76-118). These resultscoincide with previous Serological testings. The cloned NP gene has been transformed into E. coliBL21 and it could give a high expression when induced by IPTG. Western blot and ELISA assayof the purified products showed that expressed NP only reacted with NP-specific MAbs A35 and3D9, but not with G2-specific MA 3D8. The expressed NP can be used as a specific antigen forclinical diagnosis.