狂犬病病毒SRV9重排G蛋白病毒株的拯救及鉴定  

Rescue and identification of Rabies virus SRV9 G protein rearrangement virus strain

作　　者：毛丽萍 简子健[2] 翟少华[2] 潘晓梅[1] 贺笋 MAO Liping;JIAN Zijian;ZHAI Shaohua;PAN Xiaomei;HE Sun(TECON Biology Co.,Itd.,Urumqi 830011,China;College of Veterinary Medicine,Xinjiang Agricultural University,Urumqi 830052,China)

机构地区：[1]天康生物股份有限公司,乌鲁木齐830011 [2]新疆农业大学动物医学学院,乌鲁木齐830052

出　　处：《黑龙江畜牧兽医》2021年第11期17-20,26,151,152,共7页Heilongjiang Animal Science And veterinary Medicine

基　　金：国家自然科学基金项目(31360623)。

摘　　要：为了探究狂犬病病毒G蛋白的不同基因组排列位置对其毒力的影响,试验对狂犬病病毒SRV9重排G蛋白全长质粒进行测序比对及酶切鉴定,并采用反向遗传学技术将纯化后的重排全长质粒和辅助质粒共转染至BSR细胞中进行重排病毒拯救,然后采用RT-PCR扩增和间接免疫荧光方法检测盲传至第5代的重排病毒。结果表明:重排全长质粒经测序在各基因连接部位完全正确,L基因有3处碱基发生突变,转染后有少量BSR细胞产生绿色荧光;重排病毒片段经RT-PCR检测得到大小分别为1 589 bp、1 473 bp、812 bp和3 823 bp的目的片段,与预期结果一致;在共聚焦显微镜下,重排病毒能够产生大量的绿色荧光。说明利用反向遗传学技术可成功获得G蛋白重排的狂犬病病毒SRV9。In order to investigate the effect of different genomic arrangement positions of Rabies virus G protein on its virulence, the full-length recombinant plasmid of Rabies virus SRV9 rearrangement G protein was sequenced and identified by enzyme digestion. The purified full-length rearranged plasmid and its helper plasmid were co-transfected into BSR cells by reverse genetic technique to save the rearranged virus. Then the fifth generation of rearranged virus was detected by RT-PCR and indirect immuno fluorescence. The results showed that the rearranged full-length plasmid was completely correct in the connection sites of each gene. There were 3 mutations in L gene, and a small number of cells in BSR cells produced fluorescence after transfection. The rearranged virus fragments were 1 589 bp, 1 473 bp, 812 bp and 3 823 bp in size by RT-PCR, which was consistent with the expected results. Under a confocal microscope, the rearranged virus produced a large amount of green fluorescence. The results indicated that SRV9 strains of G protein rearrangement virus were successfully obtained by liposome transfection technique.

关 键 词：狂犬病病毒 G蛋白 反向遗传学 重排病毒 拯救 脂质体转染法 

分 类 号：S852.65[农业科学—基础兽医学] Q51[农业科学—兽医学]