[Objective] The study aimed at cloning PKR gene from Ctenopharyngodon idellus induced by PolyI：C in vitro,so as to provide foundation for study on the anti-virus genes of C.idellus.[Method] By referring to the PKR ge...[Objective] The study aimed at cloning PKR gene from Ctenopharyngodon idellus induced by PolyI：C in vitro,so as to provide foundation for study on the anti-virus genes of C.idellus.[Method] By referring to the PKR gene sequences of zebra fish（AJ852023.1） and Carassius auratus（AY293929.1） in Genbank,three pairs of degenerate primers were designed with 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 not from the cells treated for 12 h;in addition,the expression level increased with the processing time.Part of the amplified sequence of C.idellus shared the homology of 100% and 81.48% with the sequences of carp and zebra fish separately.[Conclusion] Part of the PKR gene sequence was cloned successfully from C.idellus.Moreover,we have proved that PolyI：C induction is effective for PKR protein expression,which will provide reference for treating viral diseases of C.idellus.展开更多
基金Supported by National Natural Science Foundation of Zhejiang Province (Y3110432 )Huzhou Teachers College Science ResearchFoundation (2010YZ48)~~
文摘[Objective] The study aimed at cloning PKR gene from Ctenopharyngodon idellus induced by PolyI：C in vitro,so as to provide foundation for study on the anti-virus genes of C.idellus.[Method] By referring to the PKR gene sequences of zebra fish（AJ852023.1） and Carassius auratus（AY293929.1） in Genbank,three pairs of degenerate primers were designed with 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 not from the cells treated for 12 h;in addition,the expression level increased with the processing time.Part of the amplified sequence of C.idellus shared the homology of 100% and 81.48% with the sequences of carp and zebra fish separately.[Conclusion] Part of the PKR gene sequence was cloned successfully from C.idellus.Moreover,we have proved that PolyI：C induction is effective for PKR protein expression,which will provide reference for treating viral diseases of C.idellus.
