| 表达H5亚型禽流感病毒HA蛋白的重组禽痘病毒的构建 |
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| 引用本文： | 焦同路,王靖飞,赵双成,王世达,常晓飞,田石,柳金雄,李雁冰,姜永萍,陈化兰.表达H5亚型禽流感病毒HA蛋白的重组禽痘病毒的构建[J].中国预防兽医学报,2012(5):341-344. |
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| 作者姓名： | 焦同路 王靖飞 赵双成 王世达 常晓飞 田石 柳金雄 李雁冰 姜永萍 陈化兰 |
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| 作者单位： | 中国农业科学院哈尔滨兽医研究所农业部动物流感重点开放实验室/兽医生物技术国家重点实验室;中国农业科学院哈尔滨兽医研究所疫病诊断与技术服务中心 |
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| 基金项目： | 哈尔滨市科技攻关计划项目(2011AA6BN019) |
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| 摘 要： | 为构建表达H5亚型禽流感病毒(AIV)A/Duck/Anhui/1/06(H5N1)(简称AH/06)HA蛋白的重组禽痘病毒,本研究利用感染-转染的方法通过转移载体pSY681-gfp-gpt将AIV株AH/06的HA基因同源重组到禽痘病毒(FPV)中,并利用gfp和gpt双重筛选标记在鸡胚成纤维细胞中经过多轮筛选,得到纯化的重组禽痘病毒(rFPV-gfp-gpt-AHHA)。通过PCR、序列测定和western blot的方法对rFPV-gfp-gpt-AHHA进行鉴定和抗原活性分析。结果表明AH/06的HA基因稳定的重组到rFPV-gfp-gpt-AHHA中,其表达的HA蛋白能够与AH/06的阳性血清反应,显示出了良好的抗原性。同时病毒的生长曲线也显示外源HA基因的插入并没有影响亲本病毒的复制。这些结果为进一步的免疫效力研究奠定了基础。
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| 关 键 词： | H5亚型禽流感病毒 重组禽痘病毒 HA基因 |
Construction of recombinant folwpox virus expressing HA protein of H5 subtype avian influenza virus |
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| JIAO Tong-lu,WANG Jing-fei,ZHAO Shuang-cheng,WANG Shi-da,CHANG Xiao-fei,TIAN Shi,LIU Jin-xiong,LI Yan-bing,JIANG Yong-ping,CHEN Hua-lan.Construction of recombinant folwpox virus expressing HA protein of H5 subtype avian influenza virus[J].Chinese Journal of Preventive Veterinary Medicine,2012(5):341-344. |
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| Authors: | JIAO Tong-lu WANG Jing-fei ZHAO Shuang-cheng WANG Shi-da CHANG Xiao-fei TIAN Shi LIU Jin-xiong LI Yan-bing JIANG Yong-ping CHEN Hua-lan |
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| Institution: | 1(1.Animal Influenza Laboratory of the Ministry Agricultural,State Key Laboratory of Veterinary Biotechnology,Harbin Veterinary Research Institute,Chinese Academy of Agricultural Sciences,Harbin 150001,China;2.Disease Diagnosis and Technical Service Center, Harbin Veterinary Research Institute,Chinese Academy of Agricultural Sciences,Harbin 150001,China) |
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| Abstract: | To construct recombinant fowlpox virus(FPV) expressing HA protein of H5 subtype avian influenza virus(AIV),the HA gene from AIV A/Duck/Anhui/1/06(H5N1) was amplified by PCR and inserted into the transfer vector pSY681-gfp-gpt of FPV.The recombinant FPV(rFPV-gfp-gpt-AHHA) was generated by transfection of the recombinant transfer vector into the quail origin FPV pre-infected chicken embryo fibroblast and purified for several cycles of selection by using Marker genes of gfp and gpt.The rFPV-gfp-gpt-AHHA was identified by PCR,sequencing and western blot.The results proved that HA gene of AIV was integrated and expressed in the recombinant FPV.One-step growth curve showed that replication of rFPV-gfp-gpt-AHHA was similar to the parent FPV.However,the protection of rFPV-gfp-gpt-AHHA needs to further evaluate in chickens against AIV infection. |
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| Keywords: | H5 subtype avian influenza virus recombinant fowlpox virus HA gene |
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