基因Ⅶ型新城疫病毒F蛋白在昆虫细胞中的表达及免疫效力研究

发布时间：2018-05-09 16:22

本文选题：新城疫病毒 + 融合蛋白F　；参考：《中国畜牧兽医》2017年06期

【摘要】：本试验旨在利用杆状病毒表达系统对基因Ⅶ型新城疫病毒(NDV)F基因进行表达研究。RT-PCR扩增F基因,将其克隆到pFastBac HT A载体中,阳性重组质粒转化DH10Bac感受态细胞,PCR鉴定获得阳性克隆,碱裂解法提取阳性质粒,转染Sf9昆虫细胞,获得含F基因的重组杆状病毒质粒,重组病毒感染Sf9细胞72h后,进行SDSPAGE电泳、间接免疫荧光和Western blotting检测。免疫SPF鸡,间接ELISA测定抗体滴度,攻毒保护试验检测重组F蛋白保护性。结果显示,F蛋白在昆虫细胞中能够特异性表达,该蛋白诱导了高滴度的NDV特异性抗体,具有良好的免疫原性;重组F蛋白免疫组攻击保护率达到90%,明显高于阴性对照组。本研究结果为NDV新型亚单位疫苗的研究奠定了基础。
[Abstract]:The aim of this experiment was to use baculovirus expression system to express gene VII (NDV) F gene and.RT-PCR amplification of F gene. The gene was cloned into pFastBac HT A vector, the positive recombinant plasmid was transformed into DH10Bac receptive cells, the positive clones were obtained by PCR identification, the positive plasmids were extracted by alkaline lysis, and Sf9 insect cells were transfected into Sf9 insect cells. Recombinant baculovirus plasmid containing F gene, after recombinant virus infection Sf9 cell 72h, SDSPAGE electrophoresis, indirect immunofluorescence and Western blotting detection. Immune SPF chicken, indirect ELISA assay antibody titer, attack protection test to detect the recombinant F protein protection. The results show that F egg white can be specifically expressed in insect cells, this protein lure. The NDV specific antibody with high titer has good immunogenicity, and the protection rate of the recombinant F protein immunization group is 90%, which is significantly higher than that of the negative control group. The results of this study lay the foundation for the study of the new subunit vaccine of NDV.

【作者单位】：徐州生物工程职业技术学院;
【基金】：徐州生物工程职业技术学院畜牧兽医品牌专业建设项目 2015年徐州市科技计划项目(KC15N0016)
【分类号】：S852.65

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