腺病毒载体介导编辑chNHE1基因对J亚群禽白血病病毒抗性研究  

Adenoviral Vector-mediated Editing of chNHE1 Gene for Avian Leukosis Virus Subgroup J Resistance Study

作　　者：王震 陈运通 魏天悦 陈洪岩[2,3] 王旭光 高玉龙[2] 夏长友[2,3] 孟庆文[2,3] WANG Zhen;CHEN Yuntong;WEI Tianyue;CHEN Hongyan;WANG Xuguang;GAO Yulong;XIA Changyou;MENG Qingwen(College of Animal Science and Technology,Xinjiang Agricultural University,Urumqi,Xinjiang 830052;State Key Laboratory for Animal Disease Control and Prevention,Harbin Veterinary Research Institute,Chinese Academy of Agricultural Sciences,Harbin,Heilongjiang 150069;National Poultry Laboratory Animal Resource Center,Heilongjiang Provincial Key Laboratory of Laboratory Animal and Comparative Medicine,Harbin,Heilongjiang 150069)

机构地区：[1]新疆农业大学动物科学学院,新疆乌鲁木齐830052 [2]中国农业科学院哈尔滨兽医研究所,动物疫病防控全国重点实验室,黑龙江哈尔滨150069 [3]黑龙江省实验动物与比较医学重点实验室,国家禽类实验动物资源库,黑龙江哈尔滨150069

出　　处：《中国家禽》2025年第2期27-33,共7页China Poultry

基　　金：国家重点研发计划项目(2022YFF0710501、2022YFF0711005);中央级公益性科研院所基本科研业务费专项(1610302022018)。

摘　　要：鸡Na^(+)/H^(+)离子交换蛋白-1(chNHE1)是J亚群禽白血病病毒(ALV-J)的细胞受体,是ALV-J感染的细胞靶点。为了探索鸡抗禽白血病的育种目标,试验构建了定点编辑chNHE1基因的CRISPR/Cas9腺病毒系统,并对编辑后细胞抗ALV-J感染能力进行评价。结果显示:使用CRISPR/Cas9腺病毒载体介导的ssODNs同源重组方案可以在chNHE1中引入突变,其中36%单克隆细胞株存在W38缺失;T37缺失的细胞对ALV-J感染具备极显著的抗性;W38缺失的细胞对ALV-J感染具有完全抗性,同时具有E35A替换、P36缺失、T37缺失的细胞对ALV-J感染也具有完全抗性。研究表明,CRISPR/Cas9腺病毒载体可以有效地编辑鸡DF-1细胞,W38缺失或E35A替换P36、T37缺失都可以使DF-1对ALV-J感染具有抗性,该方案为培育抗ALV-J感染的基因编辑鸡提供技术储备。Chicken Na^(+)/H^(+) exchanger type 1(chNHE1)is a cellular receptor for avian leukosis virus subgroup J and a cellular target of ALV-J infection.To explore the breeding goal of chicken resistance to avian leukaemia,experiment,a targeted CRISPR/Cas9 adenoviral editing system for the chNHE1 gene was constructed and the changes in cellular resistance to ALV-J after editing were evaluated.The results showed that the mutations in chNHE1 were introduced effectively by CRISPR/Cas9 adenoviral vector-mediated homologous recombination protocol of Donor,and there was 36%of monoclonal cell lines with W38 deletion.The cells with T37 deletion possessed highly significant resistance to ALV-J infection.The cells with W38 deletion and the E35A cells with modified P36 and T37 deletions were found to be completely resistant to ALV-J infection.The results of this experiment indicated that chicken DF-1 cells could be edited effectively by CRISPR/Cas9 in adenoviral vector,and either W38 deletion or E35A replacement of P36,T37 deletion could make DF-1 resistant to ALV-J infection.This protocol could be used to a technological stock for breeding chicken lines with resistance to ALV-J infection.

关 键 词：J亚群禽白血病病毒 鸡Na^(+)/H^(+)交换蛋白-1 腺病毒载体 CRISPR/Cas9 

分 类 号：S831.2[农业科学—畜牧学]