关键词: PPRV, P蛋白, 表达, 纯化, 抗体制备, 应用

Abstract: This experiment was conducted to express Peste des Petits Ruminants Virus (PPRV) phosphoprotein and prepare antiserum of phosphoprotein. For expressing the recombinant phosphoprotein in prokaryotic expression system, the recombinant plasmid pET-30a(+)-P sequenced correctly was transformed into Escherichia coli cells. Phosphoprotein was prokaryotic expressed in E. coli cell by induction of IPTG. SDS-PAGE analysis was performed to detect the recombinant protein. The highest soluble expression level of recombinant protein was obtained after being induced at 28 ℃ for 6 h and the concentration of IPTG was 0.2 mmol·μL^-1. The soluble recombinant protein was purified by Ni+ Sepharose affinity chromatograph method, and was subsequently used to immunize mouse. The anti-phosphoprotein polyclonal antibody with high sensitivity (1∶25 600) and specificity was obtained, and was used in detection of phosphoprotein eukaryotic expression in Vero. Phosphoprotein was highly expressed in E. coli, the antiserum of high titers and specificities was prepared, which lays the foundation for further research on phosphoprotein function.

Key words: PPRV, phosphoprotein, expression, purification, polyclonal antibody preparation, application