新城疫病毒感染性克隆全长cDNA转录载体的构建  

Construction of A Full-length cDNA Cloning Transcription Vector for Newcastle Disease Virus Infectious Clone

作　　者：郭承意 仝丽娜 周雷 李增魁[1] 文英[1] 刘海金[2] 杨增岐[2] 高小龙 GUO Chen-yi;TONG Li-na;ZHOU Lei;LI Zeng-kui;WEN Ying;LIU Hai-jin;YANG Zeng-qi;GAO Xiao-long(College of Agriculture and Animal Husbandry,Qinghai University,Xining,Qinghai, 810000,China;College of Veterinary Medicine,Northwest A&F University,Yangling,Shaanxi,712100,China)

机构地区：[1]青海大学农牧学院,青海西宁810000 [2]西北农林科技大学动物医学院,陕西杨凌712100

出　　处：《动物医学进展》2021年第4期22-26,共5页Progress In Veterinary Medicine

基　　金：青海省自然科学基金青年项目(2018-ZJ-952Q)。

摘　　要：以pEGFP-N1和pAdTrack-CMV质粒为模板,通过PCR扩增分别获得GFP基因序列和Kan基因序列;以胶回收的目的序列为模板,通过Overlap PCR扩增获得GFP-Kan基因序列。回收GFP-Kan基因序列,插入至pCI-neo载体上,酶切和测序鉴定正确的重组载体命名为pCI-GFP/Kan。再将设计合成的linker基因序列通过酶切、连接克隆至pCI-GFP/Kan载体上,涂布Kan抗性平板进行阳性克隆筛选,酶切和测序鉴定正确的重组载体命名为pCI-infect,即为新城疫病毒全长cDNA双启动子克隆载体。酶切和测序鉴定结果显示,pCI-infect载体构建成功。pEGFP-N1 and pAdTrack-CMV plasmids were used as templates to amplify GFP and Kan sequence by PCR,respectively.Subsequently,the GFP-Kan fusion fragment was acquired by overlap PCR with the gel-purified GFP and Kan sequences as templates.Then the gel-purified GFP-Kan fusion fragment was inserted into the pCI-neo vector.After the enzyme digestion and sequencing identification,it was named as pCI-GFP/Kan.Finally,the synthesized linker sequence was cloned into pCI-GFP/Kan plasmid by enzyme digestion and ligation.The transformation products was plated on LB agar plate(with kanamycin)for positive clones screening.After the enzyme digestion and sequencing identification,it was named as pCI-infect,namely NDV full-length cDNA cloning transcript vector.The enzyme digestion and identification results showed that pCI-infect vector was successfully constructed.

关 键 词：新城疫病毒 感染性克隆 载体 

分 类 号：S852.659.5[农业科学—基础兽医学]