摘要
[目的]克隆草鱼的PKR基因,为草鱼抗病毒基因研究奠定基础。[方法]依据GenBank上斑马鱼(AJ852023.1)和鲫鱼(AY293929.1)的PKR基因序列,利用Primer Premier 5.0软件设计了3对简并引物;采用100μg/ml PolyI:C体外分别处理草鱼肾细胞(Ctenopharyngodon indellus kidney cells,CIK)12、36、48 h,并提取处理后细胞的总RNA,逆转录后用降落PCR方法扩增这3个处理时间细胞中PKR基因。[结果]处理12 h时未扩增出PKR基因,处理36和48 h时都扩增到了PKR基因,并且表达量随处理时间的增加有所升高,扩增到的部分序列与鲫鱼和斑马鱼的该段序列同源性分别为100.00%和81.48%。[结论]试验成功获得了草鱼PKR基因的部分序列,PolyI:C高效诱导草鱼PKR蛋白表达将有助于开创治疗草鱼类病毒病的一种新思路。
[Objective]The study aimed at cloning PKR gene of Ctenopharyngodon idellus induced by PolyI： C in vitro,so as to provide foundation for study on the anti-virus genes of C.idellus.[Method]On the basis of the PKR gene sequences of zebra fish（AJ852023.1） and carp （AY293929.1） in Genbank,three pairs of degenerate primers were designed using Primer Premier 5.0 software;in vitro C.idellus kidney cells（CIK） were treated with 100 μg/ml of Poly I： C for 12,36 and 48 h,and then total RNA of the cells treated was extracted for amplifying the PKR gene by RT-PCR.[Result]The PKR gene was amplified from the cells treated with Poly I： C for 36 and 48 h.but the cells treated for 12 h;in addition,the expression level increased with the processing time.The amplified partial sequence of C.idellus shared homology of 100.00% and 81.48% with carp and zebra fish sequence separately.[Conclusion]Part of the PKR gene sequence was cloned successfully from C.idellus,PolyI： C induction was effective to PKR protein expression.
出处
《安徽农业科学》
CAS
北大核心
2011年第33期20514-20516,20529,共4页
Journal of Anhui Agricultural Sciences
基金
浙江省自然科学基金资助项目(Y3110432)
湖州师范学院校级科研资助项目
作者简介
作者简介李景芬（1977-），女，黑龙江海伦人，讲师，博士，从事动物遗传育种研究，E—mail：lijingfen@hutc．zj．cn。
