Cloning, prokaryotic expression and purification of tumor antigen gene MAGEª²3

    MA Jiaª²Hai, SUI Yanª²Fang, YE Jing, HUANG Yaª²Yu, CHEN Guangª²Sheng, SONG Hongª²Ping, ZHANG Xiuª²Min

    Department of Pathology, School of Basic Medicine, Fourth Military Medical University, Xi¬ðan 710033, China

    ¡¾Abstract¡¿ AIM: To clone human MAGEª²3 gene, to induce its prokaryotic expression and to purify the protein. METHODS: Full length cDNA of MAEGª²3 gene was amplified by RTª²PCR from human melanoma cell line. 3¡äª²terminal 324 bp segment of MAGEª²3 gene was amplified by PCR. After the gene was sequenced, it was cloned into the expression vector pGEXª²4Tª²1 to construct the expression plasmid pGEX/MAGEª²3. The DH5¦Ácontaining the expression plasmid was induced. RESULTS: The sequence of the amplified segment was identical with that reported in GenBank. The expression plasmid pGEX/MAGEª²3 was successfully constructed. The DH5¦Á containing the plasmid expressed a Mr 42 000 fusion protein after being induced by IPTG. Its purity was 89%. CONCLUSION: The full length cDNA of MAEGª²3 gene was cloned. The 108 aa segment of MAGEª²3 encoded by 3¡äª²terminal 324 bp of gene is expressed and purified successfully, which lays a foundation for the application of the protein in tumor immunotherapy. ......