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浙江大学学报（农业与生命科学版）
浙江大学学报(农业与生命科学版)  2011, Vol. 37 Issue (5): 473-478    DOI: 10.3785/j.issn.1008-9209.2011.05.001
生物科学与技术     
稳定表达传染性法氏囊病病毒VP2蛋白突变体细胞系的建立
孙静,魏永伟,章晓栋,于涟
浙江大学动物科学学院 浙江省动物预防医学重点实验室,浙江杭州 310058
Construction of cell line stably expressing IBDV VP2 protein mutants
SUN Jing,WEI Yong-wei,ZHANG Xiao-dong,YU Lian
Zhejiang Province Key Lab of Preventive Veterinary Medicine , College of Animal Sciences , Zhejiang University , Hangzhou 310058 , China
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摘要： 根据传染性法氏囊病病毒( IBDV) HZ２ 株VP２ 基因序列设计１ 对引物,将该基因成功克隆到真核表达载体pDsRed２‐N１ 中,构建重组质粒pDsRed２‐N１‐VP２(简称PVP２) ;以PVP２ 作为过渡载体,运用PCR 定点突变技术,对第２７９ 位点和第２８４ 位点的氨基酸残基分别或同时进行突变,共构建P２７９ 、P２８４ 和PDA ３ 株突变体;在此基础上构建能稳定表达HZ２ 株IBDV 及其突变体VP２ 蛋白的Vero 细胞系,间接免疫荧光试验检测表明,Vero 细胞系中IBDV VP２ 蛋白已成功表达.这为进一步深入研究IBDV 毒力及抗原变异致病机制奠定了基础;此外,利用定点突变技术改造VP２ 蛋白,建立不同毒力的突变体,在探求高效、廉价的IBDV DNA 疫苗上也有广阔的应用前景和重要意义.
Abstract: A couple of primers was designed on the base of VP２gene sequence of infectious bursal disease virus (IBDV) HZ２strain , and the VP２gene was successfully cloned into eukaryotic expression vector pDsRed２‐N１ . Then , the recombinant plasmid pDsRed２‐N１‐VP２ was constructed . The ２７９site and ２８４site on VP２ protein were mutated singly or homeochronously through site‐directed mutagenesis on theconstructed vector pDsRed２‐N１‐VP２ . By this way , three mutants of P２７９ , P２８４ and PDA were constructed in next step . The Vero cell line stably expressing VP２ protein and this mutant were also constructed successfully on the base of P２７９ , P２８４ and PDA by indirect immunofluorescence test . Therecombination plasmid and cell line constructed lay the foundation on the deep research about the role and mechanism of ２７９ site and ２８４ site in virulence change and antigenic variation of IBDV , which also play a significant role in exploring more efficient and cheaper IBDV DNA vaccination .
出版日期: 2011-09-20
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孙静
魏永伟
章晓栋
于涟

引用本文:

孙静,魏永伟,章晓栋,于涟. 稳定表达传染性法氏囊病病毒VP2蛋白突变体细胞系的建立[J]. 浙江大学学报(农业与生命科学版), 2011, 37(5): 473-478.

SUN Jing,WEI Yong-wei,ZHANG Xiao-dong,YU Lian. Construction of cell line stably expressing IBDV VP2 protein mutants. Journal of Zhejiang University: Agric. & Life Sci., 2011, 37(5): 473-478.

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http://www.zjujournals.com/agr/CN/10.3785/j.issn.1008-9209.2011.05.001    或    http://www.zjujournals.com/agr/CN/Y2011/V37/I5/473

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