Expression Of Truncated Glycoprotein Of Vesicular Stomatitis Virus In E.coli And The Research Of Diagnostic Technique

Vesicular stomatitis (VS) is classified as a list A disease by the Office International des Epizooties (OIE) and a second class disease in China's import and export inspection and quarantine of animal and animal products. Following China entered into WTO, the international trade for animal and animal products has been increasing, though China has no news about prevalence of VS at present. It is necessary for China to develop research on this disease to prevent it spreading to China and protect the health development of our animal husbandry.VS is an acute and high contagious disease for various species of mammals caused by vesicular stomatitis virus (VSV) of the family Rhabdovi-ridae, genus Vesiculovirus. VSV virion is composed of five different proteins, nucleocapsid protein (N), phosphoprotein (P), matrix protein (M), glycoprotein(G), and polymerase protein (L). N and G protein have got great significance for immunity and diagnosis of the five proteins. N protein encoded by N gene can stimulate the nonneutralization antibody, which is responsible for group of VSV. G protein encoded by G gene appears is the major antigenic determinant and is responsible for neutralization of infectivity by antibody and for type, subtype and even strain specificity of virus.In this study, the G gene of vesicular stomatitis virus Indiana (VSV-IND) without signal and transmembrane region was amplified by PCR. The DNA sequenced EcoR I and Xho I restriced core fragment was cloned in a frame downstream of T7 promoter in the pET-30c vector, then the recombinant plasmids pET-30c-vsvG were transformed into E.coli BL21(DE3). The recombinant fusion protein was highly expressed in E.coli BL21 four hours later induced by 1 mmol/L IPTG, and the molecular weight of the fusion protein was about 57 ku, subsequently examined by SDS-PAGE. Western-blot showed that the expressed recombinant protein could be recognized by VSV-IND positive serum.The recombinant protein containing a six-Histidine tag aggregated into the inclusion body could be purified by Ni-column, which was as the coated antigen after purified in the established indirect ELISA. The result showed that the recombinant protein purified had good antigenicity and type specificity. Therefore, the recombinant protein will replace the...