Expression And Purification Of The Getah Virus E2 Protein And Initial Establishment Of RT-PCR For The Detection Of GETV

Getah virus belong to the Togaviridae family of the Alphavirus, it can spread the disease through the way of mosquitoes bites vertebrate. When GETV prevalent in pigs,it is harm to the pig-breeding industry,the pathogenicity of GETV has been shown in pregnant sows and newborn piglets. GETV is one of the causes of reproductive disorders in pregnant sows and fever, rash, edema of the hind legs, depression and diarrhea in newborn piglets. There is still noteffective drugs and vaccine forGETV disease. Therefore,it is very important to establish a specific and accurate detection method for the detection of GETV.The E2 gene of GETV strain (SH05-6) was optimized according to the complete genome sequence of structural protein E2 published in GenBank (EU015066) and synthesized artificially in this experiment. The synthesized E2 gene was cloned into the expression vector pCold I for generation of a recombinant plasmid pCold I-E2.The plasmid pCold I-E2 was induced with IPTG in low temperature and the expression of the targeted protein was detected by SDS-PAGE and Western Blot. Res μ Its showed that the expressed protein was a a relative molecular mass of 46KD and in line with expectations. Approximately 80% of total proteins of E.coli was the expressed E2 protein. The E2 gene was highly expressed in the E.coli system by low temperature. The condition of protein purification was optimized and innovated the method of protein purification, ultimately the high purified E2 proteins was obtained.RT-PCR for the detection of GETV was established through primer design by E2 gene and optimized the reaction conditions. The res μ It revealed that no GETV can be found in the mosquitoes from the regions of Shanghai when used this RT-PCR to detect GETV. This experimental will promote the research and development of rapid diagnostic technology for GETV detection.