芜菁皱缩病毒P38蛋白的原核表达及抗血清的制备  

Prokaryotic expression of P38 fromTurnip Crinkle Virus and preparation of its polyclonal antibody

作　　者：张静[1] 朱峰[1] 陈应娟[1] 邓星光 刘健[1] 林宏辉[1] 席德慧[1] 

机构地区：[1]四川大学生命科学学院生物资源与生态环境教育部国家重点实验室,成都610064

出　　处：《四川大学学报（自然科学版）》2015年第2期441-445,共5页Journal of Sichuan University(Natural Science Edition)

基　　金：国家自然科学基金(31171835;31270290);四川省应用基础项目(2012JY0078);中央高校基本科研业务费(2011SCU04B34)

摘　　要：从感染芜菁皱缩病毒(TCV)的拟南芥中提取总RNA,通过反转录PCR(RT-PCR)扩增得到TCV的外壳蛋白基因P38.在目的基因ORF框两边引入酶切位点BamHⅠ和SacⅠ,并连接到表达载体pET30-a上,导入大肠杆菌BL21(DE3)中,37℃经0.5mM/L IPTG诱导6h后获得了分子量约为48kD的融合蛋白.经Ni-NTA纯化目的蛋白,注射大白兔制备抗血清.Western blot结果表明制备好的抗血清能成功检测感染TCV的拟南芥.The nucleotide sequence of the coat protein（P38）gene of Turnip crinkle virus was obtained using RT-PCR fromArabidopsis thalianainfected with TCV.Restriction enzyme cutting sites of BamHⅠ and SacⅠ were introduced to the ORF fragment of P38.Then the fragment was linked together with PET30-a and transformed into E.coli BL21（DE3）.The target fusion protein（48kD）was induced by0.5mM/L IPTG at 37℃for 6h.The expressed protein was purified by Ni-NTA and anti-P38 polyclonal antibody was prepared using rabbit.Western blot results showed that the antiserum could be used to detect the expression of P38 in infected Arabidopsis thaliana.

关 键 词：芜菁皱缩病毒 外壳蛋白 原核表达 抗血清制备 

分 类 号：Q78[生物学—分子生物学]