甘薯羽状斑驳病毒和褪绿矮化病毒重组双CP融合基因的原核表达和抗血清的制备

Prokaryotic Expression of Recombinant Double CP Fused Gene of Sweet potato feathery mottle virus and Sweet potato chlorotic stunt virus and Preparation of Antiserum

由褪绿矮化病毒(SPCSV)和羽状斑驳病毒(SPFMV)协生共侵染引起病毒病(SPVD)是甘薯最严重的病害之一。为了建立一种高效的甘薯病毒病SPVD检测体系,本研究构建了SPCSV和SPFMV重组双CP基因的原核表达载体pET22b-SPVD-CP,该重组菌在20℃条件下经IPTG诱导,获得了68 kDa的融合蛋白(重组双CP)。利用高纯度重组双CP为抗原免疫家兔,获得了SPVD的抗血清,间接ELISA检测显示抗体稀释度在1∶512 K的范围内,制备的抗体与抗原具有较强的结合活性。本研究结果为利用重组CP作抗原大量制备SPVD抗体奠定了基础。

SPVD is one of the most serious viruses in sweetpotato,which is caused by co-infection of sweet potato feathery mottle virus(SPFMV)and sweet potato chlorotic stunt virus(SPCSV).In order to establish an efficient detection system for virus disease SPVD of sweet potato,a prokaryotic expression vector harboring CP genes of both SPCSV virus and SPFMV virus was constructed and named pET22b-SPVD-CP.After induction of the recombinant bacteria with IPTG at 20℃,the fusion gene was successfully expressed and 68 kDa fusion protein(the recombinant CP)was obtained.The purified recombinant CP was used as antigen to immune rabbits and antiserum specific to SPVD was obtained,its titer was 1∶516K in indirect detection.This research has laid the foundation for preparation of antiserum against SPVD by using recombinant CP as antigen in a large scale.