传染性法氏囊病病毒C4毒株VP2基因的原核表达及生物信息学分析

发布时间：2018-03-21 13:30

本文选题：传染性法氏囊病病毒(IBDV)　切入点：VP　出处：《中国畜牧兽医》2017年12期 　论文类型：期刊论文

【摘要】：试验旨在研究一株传染性法氏囊病病毒(IBDV)河南分离株的毒力特征及其与VP2氨基酸序列特征的关系。通过提取IBDV C4株RNA,利用RT-PCR扩增其VP2基因,与其他不同毒力IBDV毒株进行核苷酸及推导的氨基酸序列比对分析,同时使用pET-32a(+)原核表达载体表达VP2基因,用SDS-PAGE和Western blotting检测重组VP2蛋白的表达。结果显示,扩增的IBDV C4株的VP2基因序列在进化关系上属于超强毒力IBDV(vvIBDV)分类,与选取的vvIBDV毒株代表毒株核苷酸序列同源性在98.1%~98.7%之间,其七肽区为S-W-S-AS-G-S(第326—332位氨基酸)符合超强毒株特征,且222(A)、256(I)、294(I)和299(S)位氨基酸与超强毒力毒株的4个特征性氨基酸一致;但IBDV C4毒株的VP2蛋白氨基酸序列与超强毒力毒株代表毒株UK661相比,201(D/G)、281(G/R)、313(V/A)位氨基酸不同,其中281位氨基酸的改变处于279—290的小亲水区内,与病毒抗原性有关;构建的pET-32a(+)-VP2原核表达载体在大肠杆菌BL21感受态细胞上表达出分子质量约67ku的重组VP2蛋白,为进一步比较201(G)、281(R)、313(A)位氨基酸差异导致的抗原特性改变提供了研究基础。本试验结果表明,IBDV C4株VP2基因与vvIBDV毒株VP2基因的主要特性一致,但也有3处氨基酸与代表毒株UK661存在差异,这些改变可能与中国IBDV毒株毒力的进化有关。
[Abstract]:The purpose of this study was to study the virulence of a strain of infectious bursal disease virus (IBDV) from Henan province and its relationship with the amino acid sequence of VP2. The VP2 gene of IBDV C4 strain was amplified by RT-PCR. Nucleotides and deduced amino acid sequences were compared with other virulent IBDV strains. The VP2 gene was expressed by pET-32a () prokaryotic expression vector, and the expression of recombinant VP2 protein was detected by SDS-PAGE and Western blotting. The VP2 gene sequence of the amplified IBDV C4 strain belongs to the highly virulent IBDVV vIBDVV classification in the evolutionary relationship. The nucleotide sequence homology with the selected vvIBDV strain is 98.1% and 98.7% respectively, and the heptapeptide region is S-W-S-AS-G-S (326-332 amino acid) in accordance with the characteristics of the supervirulent strain. The amino acid sequence of VP2 protein of IBDV C4 strain was different from that of UK661 strain, and the amino acid sequence of VP2 protein of strain C 4 was different from that of strain UK661, and the amino acid sequence of VP2 protein of strain C 4 was different from that of strain UK661, and the amino acid sequence of VP2 protein of strain C 4 was different from that of strain UK661, and the amino acid sequence of VP2 protein of strain C 4 was different from that of strain UK661, and the amino acid sequence of VP2 protein of strain C 4 was different from that of strain UK661, and the amino acid sequence of VP2 protein of strain C 4 was different from that of strain UK661. The alteration of 281 amino acids was located in the small hydrophilic region of 279-290, which was related to the antigenicity of the virus. The constructed pET-32a (pET-32a) prokaryotic expression vector expressed a recombinant VP2 protein with a molecular weight of about 67 ku in Escherichia coli BL21 receptive cells. In order to further compare the changes of antigenic characteristics caused by the amino acid difference of RDV 313A, the results showed that the main characteristics of VP2 gene and VP2 gene of IBDV C4 strain were consistent with those of vvIBDV strain VP2, but there were also three amino acids differences between the three amino acids and the representative strain UK661. These changes may be related to the evolution of virulence of IBDV strains in China.
【作者单位】：河南农业大学牧医工程学院;
【基金】：国家十三五科技专项(2017YFD0500701)
【分类号】：S852.65

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