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| 猪圆环病毒2型ORF2基因的原核表达及其免疫原性检测 | |
| 引用本文： | 梁晓艳,李玉林,李兆学,王继美,贾丽锋,周贺娟,王云龙.猪圆环病毒2型ORF2基因的原核表达及其免疫原性检测[J].动物医学进展,2010,31(9). |
| 作者姓名： | 梁晓艳 李玉林 李兆学 王继美 贾丽锋 周贺娟 王云龙 |
| 作者单位： | 1. 郑州大学,河南郑州,450002 2. 河南省生物工程技术研究中心,河南郑州,450002 3. 河南工业大学,河南郑州,450002 4. 郑州大学,河南郑州,450002;河南省生物工程技术研究中心,河南郑州,450002 |
| 摘 要： | 以病猪脾脏组织为材料,提取总RNA,通过RT-PCR得到579 bp的猪圆环病毒2型(PCV-2)/ORF2片段,与原核表达载体pET32a重组,通过菌落PCR鉴定、双酶切鉴定及测序鉴定后,证实重组质粒pET32a/ORF2构建成功,将其转化入表达菌Balgold,通过IPTG诱导表达,利用Ni-NTA亲和层析纯化目的产物,Western blot验证其免疫原性,重组蛋白免疫Balb/c小鼠,检测小鼠体内中和抗体滴度。获得的重组蛋白包涵体的形式出现,表达量占菌体总蛋白的30%,变性纯化后纯度达90%以上,Western blot证实重组蛋白能与PCV-2阳性血清发生反应,免疫Balb/c小鼠45 d后中和抗体滴度达到1∶22。 |
| 关 键 词： | 猪圆环病毒 原核表达 免疫原性 |
Prokaryotic Expression and Purification of PCV-2 ORF2 Gene and Its Immunogenicity Detection |
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| LIANG Xiao-yan,LI Yu-lin,LI Zhao-xue,WANG Ji-mei,JIA Li-feng,ZHOU He-juan,WANG Yun-long.Prokaryotic Expression and Purification of PCV-2 ORF2 Gene and Its Immunogenicity Detection[J].Progress In Veterinary Medicine,2010,31(9). | |
| Authors: | LIANG Xiao-yan LI Yu-lin LI Zhao-xue WANG Ji-mei JIA Li-feng ZHOU He-juan WANG Yun-long |
| Institution: | LIANG Xiao-yan1,LI Yu-lin2,LI Zhao-xue3,WANG Ji-mei2,JIA Li-feng1,ZHOU He-juan1,WANG Yun-long1,2(1.Zhengzhou University,Zhengzhou,Henan,450002,China,2.Henan Biotechnology Research Center,3.Henan University of Technology,China) |
| Abstract: | Pigs spleen was used for the extraction of total RNA,and the 579 bp fragment of PCV-2/ORF2 cDNA was amplified through RT-PCR method.The fragment was cloned into prokaryotic expression vector pET32a and transformed into Balgold.After the recombinant plasmid pET32a/ORF2 was confirmed by PCR,restriction enzyme digestion analysis and DNA sequencing,it was transduced by IPTG,the recombinant protein was purified by Ni-NTA affinity chromatography and identified by Western blot,then it was used to immunize Balb/c m... |
| Keywords: | ORF2 |
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