新城疫病毒mRNA疫苗的构建及鉴定  

Construction and identification of Newcastle disease virus mRNA vaccine

作　　者：崔苗苗 陈凯楠 李金斗 丁佳欣 陈铭桦 冯嘉轩 周铁忠 舒秀伟 刘晓雷 丁壮 CUI Miaomiao;CHEN Kainan;LI Jindou;DING Jiaxin;CHEN Minghua;FENG Jiaxuan;ZHOU Tiezhong;SHU Xiuwei;LIU Xiaolei;DING Zhuang(Key Laboratory for Zoonosis Research of Ministry of Education,College of Veterinay Medicine,Jilin University,Changchun 130062,China;Key Laboratory of Liaoning Province,College of Animal Husbandry and Veter-inary Medicine,Jinzhou Medical University,Jinzhou,Liaoning 121000,China;Liaoning Yikang Biotechnology Co.,Ltd.,Liaoyang,Liaoning 111000,China)

机构地区：[1]吉林大学动物医学学院人兽共患病研究教育部重点实验室,吉林长春130062 [2]锦州医科大学畜牧兽医学院辽宁省重点实验室,辽宁锦州121000 [3]辽宁益康生物股份有限公司,辽宁辽阳111000

出　　处：《中国兽医学报》2023年第12期2480-2485,共6页Chinese Journal of Veterinary Science

基　　金：国家自然科学基金资助项目(32072860);吉林省科技厅重点研发计划资助项目(20230202082NC,20200402037NC);辽宁省科技厅重点研发资助项目(2021JH2/10200009);长春市科技局重点研发计划资助项目(21ZGN17);辽宁省民生科技计划资助项目(2021JH2/10200009)。

摘　　要：信使RNA(messenger RNA,mRNA)的核酸疫苗是近年来兴起的一种新型疫苗技术。新城疫病毒(Newcastle disease virus,NDV)的血凝素-神经氨酸酶(HN)是NDV的主要保护性抗原,可诱导机体产生中和抗体,是新城疫(Newcastle disease,ND)新型疫苗研发的重要靶点。本研究将NDV HN基因连接到含有T7启动子的载体中,并在其5′UTR和3′UTR分别引入人-β球蛋白的非编码区域以增强转录的mRNA稳定性,经质粒线性化、体外转录、cap加帽处理、mRNA纯化制备NDV HN-mRNA,将其转染至HEK-293T细胞中验证NDV HN-mRNA的表达效果,Western blot和免疫荧光试验结果表明构建mRNA候选疫苗可在HEK-293T细胞中高效表达抗原蛋白,为ND新型疫苗的研发提供新的思路。The nucleic acid vaccine of the messenger RNA(mRNA)is a new type of vaccine technology that has emerged in recent years.Newcastle disease virus(NDV)hemoglobin-neuraminidase(HN)is the main protective antigen of the Newcastle disease virus and an important target for new types of vaccines for Newcastle disease(ND).It can induce the body to produce neutral antibodies.Therefore,this study will clone the NDV HN gene to the carrier containing the T7 promoter and introduce the non-encoded area of theβglobulin in its 5'UTR and 3'UTR to enhance the mRNA stability of the transcription.NDV HN mRNA was prepared by plasmid linearization,in vitro transcription,cap processing,and mRNA purification,then it was transfected into HEK-293T cells to verify the expression effect of NDV HN mRNA.The results of Western blot and immune fluorescence showed that the mRNA candidate vaccine can highly express antigen protein in HEK-293 Tcells,providing new ideas for the research and development of novel ND vaccines.

关 键 词：新城疫病毒 HN蛋白 体外转录 mRNA疫苗 

分 类 号：S852.65[农业科学—基础兽医学]