Expression of a human TNF-α mutant in E. coli and aeasurement of its ability to induce apoptosis

Using PCR technique, a Tumor Necrosis Factor-α(TNF-α) mutant was obtained by altering the Gly on 153 site to Leu. After the amplified hTNF gene fragment was cloned into expressing vector pQE30, the E. coli transformed by the recombinant plasmid was induced by IPTG. SDS-PAGE assay proved that hTNF-αprotein expressed in E. coli reached a high level production. The protein was collected and highly purified through Ni-NTA-agarose. Westernblot showed that the protein could react strongly with specific anti-TNF serum. The ability of hTNF protein to induce apoptosis was examined using fluorescence staining method.