口蹄疫病毒结构蛋白VP1容忍外源标签的能力

发布时间：2018-06-07 00:19

本文选题：口蹄疫病毒 + 结构蛋白　；参考：《微生物学报》2017年05期

【摘要】：【目的】利用口蹄疫病毒的反向遗传操作技术,构建含不同外源标签口蹄疫病毒的全长克隆,鉴定口蹄疫病毒结构蛋白VP1容忍不同外源标签的能力。【方法】通过融合PCR技术,在FMDV O/HN/93全长感染性克隆的VP1 G-H环分别引入V5、TC12、KT3、3FLAG外源标签,构建全长质粒。全长质粒经Not I线化后转染表达T7 RNA聚合酶的稳定细胞,拯救重组病毒。RT-PCR、序列测定、间接免疫荧光鉴定病毒,噬斑和一步生长曲线分析重组病毒的生物学特性。【结果】成功拯救到表达V5或KT3表位标签的重组病毒,未能拯救到表达TC12或3×FLAG的重组病毒。V5和KT3表位标签的插入均影响了口蹄疫病毒的复制能力。【结论】重组口蹄疫病毒的成功拯救为未来标记疫苗以及口蹄疫病毒作为表达载体等的研究奠定了基础。
[Abstract]:[Objective] to construct a full-length clone of foot-and-mouth disease virus containing foot-and-mouth disease virus (FMDV), and identify the ability of FMD VP1 to tolerate different external labels. [Methods] V5, TC12, KT3,3FL were introduced into the VP1 G-H ring of FMDV O/HN/93 full-length infected clone by the fusion of PCR technology. AG extraneous label, construct full length plasmid. The whole plasmids were transfected with T7 RNA polymerase after Not I linearization. The recombinant virus.RT-PCR, sequencing, indirect immunofluorescence identification virus, plaque and one step growth curve were used to analyze the biological characteristics of recombinant virus. [results] the weight of V5 or KT3 epitopes was successfully saved. The failure to save the expression of TC12 or 3 * FLAG recombinant virus.V5 and KT3 epitopes all affect the replication ability of foot-and-mouth disease virus. [Conclusion] the successful rescue of FMD has laid the foundation for the future study of labeled vaccine and foot-and-mouth disease virus as an expression vector.
【作者单位】：中国农业科学院兰州兽医研究所家畜疫病病原生物学国家重点实验室国家口蹄疫参考实验室;
【基金】：甘肃省农业生物技术研究与应用开发项目(GNSW-2014-22) 甘肃省青年科技基金计划(1606RJYA256)~~
【分类号】：S852.65

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