Molecular Cloning, Expression And Identification Of GP50 Gene From Cysticercus Cellulosae In Vitro

Objective: To genernate a quick and accurate detection of Cysticercosis by use of recombinant protein, GP50 was amplified from Cysticercus cellulosae and subcoloned into the expression vector pBK-CMV. Methods: The specific primers were designed and synthesized. The DNA fragment encoding GP50 was amplified by PCR from cDNA of C.cellulosae. The PCR products were cloned into pGEM-T vector and recombinants were screened by PCR, risfriction digestion followed by sequencing. Then the GP50 was subcloned into pBK-CMV expression vector and the recombinant plasmid was transformed into E. coli BL21,IPTG was added to induce fusion expression of fused GP50. The expression was identified by Western blot. Results: The size of pGEM-T-GP50,pBK-CMV-GP50 digested with BamH I and Xho I was identical to the length of the PCR generated products. DNA sequencing indicated that the GP50 gene has an open reading frame of 897bp,which encodes 298 amino acids with an approximate molecular weight of 33kDa and has high homology with the GP50 gene submitted to GenBank (AY212945). As consequence GP50 gene was successfully cloned and subcloned into the vectors. With the positive clones induced by IPTG, a fusion protein was expressed in E.coli BL21. The fusion protein could be recognized by the sera of patients infected with Cysticercus cellulosae.