摘要
本研究以水稻幼苗为材料,通过RT-PCR扩增得到1.1kb的编码PDK2(登录号:AK100033)基因的片段,成功构建重组表达载体pET-23d-PDK2并转化到大肠杆菌BL21(DE3)中。研究结果表明,重组蛋白以包涵体的形式存在,且PDK2的最佳表达条件为:28℃,120r/min,0.5mmol/L IPTG诱导60min。经纯化后免疫新西兰大白兔以制备PDK2多克隆抗体。Western blot检测表明该抗体的特异性和效价较好,这为进一步研究水稻PDK2的生物学功能奠定了基础。
In this paper,we cloned a 1.1 kb fragment encoding PDK2 from rice seedlings by using RT-PCR ap-proach,and successfully built the recombinant expression construct pET-23d-PDK2,then transformed it into E.coli BL21(DE3).Experiments showed that the recombinant protein PDK2 was expressed in the form of inclusion bodies,and the optimal expression condition for PDK2 was found to be as follows:28℃,120 r/min,0.5 mmol/L IPTG treated for 60 min.Subsequently,the recombinant protein was injected into a New Zealand white rabbit for polyclonal antibody preparation after purification.Western blot analysis indicated that the polyclonal antibody owned relatively high specificity and titer,which would lay foundations for biological function research of rice PDK2.
出处
《基因组学与应用生物学》
CAS
CSCD
北大核心
2009年第6期1039-1042,共4页
Genomics and Applied Biology
基金
广东省自然科学基金(0909037)资助
关键词
PDK2
原核表达
多克隆抗体
水稻
PDK2,Prokaryotic expression,Polyclonal antibody,Rice
