重组甲鱼载脂蛋白B的研制及其阻断STSSSV感染的活性初探
PDF下载 (314)许丽华,顾叶华,陈梅娟,李登峰.重组甲鱼载脂蛋白B的研制及其阻断STSSSV感染的活性初探[J].宁波大学学报(理工版),2020,33(2):7-12.DOI:
XU Lihua,GU Yehua,CHEN Meijuan,LI Dengfeng.Preparation and STSSSV infection blocking assay of recombinant PApoB[J].Journal of Ningbo University(Natural Science & Engineering Edition),2020,33(2):7-12.DOI:
| Title: | Preparation and STSSSV infection blocking assay of recombinant PApoB |
| 作者: | 许丽华, 顾叶华, 陈梅娟, 李登峰 |
| Author(s): | XU Lihua, GU Yehua, CHEN Meijuan, LI Dengfeng |
| 关键词: | 甲鱼; 载脂蛋白B; 原核表达; 感染阻断 |
| Keywords: | STSSSV; soft-shelled turtle; apolipoprotein B; prokaryotic expression; blocking infection |
| 分类号: | S948 |
| 文献标识码: | A |
| 摘要: | 为了制备甲鱼(Pelodiscus sinensis)载脂蛋白B (Pelodiscus sinensis Apolipoprotein B, PApoB) 基因的原核表达产物, 并探索其是否具有阻断甲鱼系统性败血症球状病毒(Soft-shelled Turtle Systemic Septicemia Spherical Virus, STSSSV)感染的功能, 为最终控制和防治甲鱼系统性败血症疾病奠定基础, 构建了PApoB的原核表达系统, 纯化了重组PApoB, 并将其用于阻断STSSSV感染的分析: 根据甲鱼PApoB的基因序列, 设计引物, 扩增其第9932-11209bp(3294-3696 aa)区段, 将其连接至表达载体pET-28a(+)中, 转化大肠杆菌Rosetta株, 经硫代半乳糖苷(IPTG)诱导表达, 并用亲和层析法纯化获得了重组PApoB; 将FITC标记的STSSSV与PApoB孵育后对甲鱼细胞进行攻毒, 显微镜下观察, 结果PApoB添加组的病毒荧光信号明显低于非添加组, 表明该重组蛋白可以阻断STSSSV的感染. |
| Abstract: | The current study was aimed to prepare the recombinant protein of Pelodiscus sinensis Apolipoprotein B (PApoB) and examine whether the recombinant PApoB has the function of blocking the infection of soft- shelled turtle (Pelodiscus sinensis) systemic septicemia spherical virus (STSSSV) Primers were designed referring to PApoB gene to amplify a 1 277 bp fragment. The fragment was ligated into the expression vector pET-28a(+). Subsequently, the recombinant plasmid was transformed into Escherichia coli Rosetta and was induced by the thiogalactoside (IPTG). PApoB was purified by affinity chromatography. Following the incubation of FITC-labeled STSSSV and recombinant PApoB, the soft-shelled turtle cells were challenged and observed under fluorescence microscope. Results showed that the fluorescence signal of the virus in the PApoB- supplemented groups was significantly lower than that of the non-PApoB-addition groups. The results indicate that the recombinant protein was effective to inhibit the infection of STSSSV. |
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| 备注/Memo: | 收稿日期: 2019－01－23. 宁波大学学报（理工版）网址: http://journallg.nbu.edu.cn/ 基金项目: 国家重点研发计划（2018YFA0903000）; 浙江省公益性技术应用研究计划（2011C32004）; 宁波市农业攻关项目（2011C10004）. 宁波大学学报（理工版）网址：http://journallg.nbu.edu.cn/第一作者: 许丽华（1995－）, 女, 安徽铜陵人, 在读硕士研究生, 主要研究方向: 水产动物病害防治. E-mail: 1136912634@qq.com *通信作者: 李登峰（1975－）, 女, 湖南双峰人, 副教授, 主要研究方向: 水产动物病害防治. E-mail: lidengfeng@nbu.edu.cn |
