Cloning Of σC,σB Gene Of Avian Reovirus And Their Prokaryotic Expression

Avian reovirus(ARV) is an important cause of disease in poultry. In particular,reovirus-induced arthritis,chronic respiratory diseases, and malabsorption syndrome cause considerable economic losses.The σ C encoded by the ARV S1 gene is epitoped of 326 amino acids and possesses both broadly-specific epitopes .The σ B encoded by the ARV S3 gene is epitoped of 368 amino acids and carries group-specific neutralizing epitopes.Both proteins are also involved in the induction of cell fusion and may play an important role in viral pathogenesis , They all possess group-specific neutralizing epitopes which have relationship with emerging of neutralizing antibody.The σ C and σ B gene of ARV were successfully amplified by Reverse Transcription-Polymerase Chain Reaction in one step using designed primers and were cloned and sequenced.Their sequences of cDNA and amino acids were analyzed with computer software on molecular biology.Based on the results of restriction enzyme map analysis,expression primers were desigened,Cloned σ C and σ B gene were inserted into the bacterial plasmids pGEX-4T-1.The recombinant plasmids containing o C or σ B were identified by restriction enzyme analysis and PCR methods.The recombinant fusion protein was highly expressed in the form of inclusion bodies in E.coli BL21 induced by IPTG Molecular weight of fusion protein σ C is about 60KDa and fusion protein o B is 64KDa,they are identical with that of expected..Western-blot analysis with ARV -positive serum showed the fusion proteins had good specificity.The purifed σ C and σ B protein were identified using ELISA method.The results show that these purifed proteins have good activity.In this study, the purified protein σ C and σ B also can be tested for their immunogenic feature and their function in the pathogenesis.