Cloning and Expression of Extracellular Domain of Prostate Specific Membrane Antigen in Escherichia coli and Preparation of Polyclonal Antibody
人前列腺特异膜抗原膜外区基因的克隆表达及抗体的制备

Human Prostate Specific Membrane Antigen(PSMA) cDNA was amplified using total RNA extracted from prostate carcinoma tissue by RT-PCR. The cDNA fragment of extracellular domain of PSMA(edPSMA) gene was amplified by PCR and cloned into expression vector pMAL-c2x. Sequence analysis of both PSMA and edPSMA revealed identity to the GenBank reported. The edPSMA was expressed in E. coli as part of a fusion protein with MBP as the induction of IPTG. Western blot analysis showed the recombinant protein could react with PSMA monocloned antibodies 4G5. MBP-edPSMA fusion protein were purified by amylose resin affinity chromatography and showed to be homogeneity in SDS-PAGE(120 kD). BALB/C mice were immunized with the purified protein for the preparation of polyclonal antibody. The polyclonal antibody, which had a title of 1:12,800, were indicated the specificity to prostate tissue.