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Asian Journal of Animal and Veterinary Advances
Year: 2012  |  Volume: 7  |  Issue: 12  |  Page No.: 1301 - 1311

Cloning, Expression and Purification of Recombinant Envelope Protein VP36A of White Spot Syndrome Virus

Xinyan Leng and Rongmei Fei    

Abstract: White spot syndrome is a viral infection of penaeid shrimps which is highly infectious and lethal, terminating shrimps rapidly. Outbreaks of this disease can wipe out entire shrimp populations within a few days. To reduce the mortality of shrimps and increase the production of marine industry, the research focused on the changes on the virus’s proteins. The research is aimed at the molecular reconstruction of one of the important proteins from the White Spot Syndrome Virus (WSSV). By exploring the unique DNA sequences as well as the corresponding protein, the initial target shall be achieved soon. According to the published gene sequence of White Spot Syndrome Virus (WSSV), a pair of specific primers were designed. Using an isolate of WSSV collected in Ganyu, Jiangsu Province, China, as template, a gene fragment of VP36A was amplified by Polymerase Chain Reaction (PCR). The PCR product was firstly TA-cloned into a pMD-18T vector and then inserted into a pET-28a plasmid to create a recombinant plasmid and finally transformed into the host strain BL21. The recombinant protein was expressed in the form of inclusion bodies with 1.0 mmol L-1 Isopropyl-β-D-thiogalactopyranoside(IPTG) at 37°C. According to the results of an Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis (SDS-PAGE), the recombinant protein had the expected size of 36 kDa. The purified recombinant protein was tested in a Western blot to confirm that the target protein had been successfully expressed. This showed that the special DNA in WSSV can be replicated and the protein can be expressed by a very cheap and sufficient way in a very limiting time, which will be utilized in the further experiments related to the functional identification of protein (VP36A).

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