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| Efficient Preparation of TNFα Derivatives TRSP10 and Preliminary Study of Its Inhibitory Effect on Prostate Cancer DU145 Cells |
| FANG Shi-xiong, MA Yi, SHEN Shu-tao, ZHAO Shao-jun, HONG An |
| Department of Cell Biology of Jinan University, Institute of Biological Medicine of Jinan University, Guangdong Provincial Key Laboratory of Bioengineering Medicine, National Engineering Research Center of Genetic Medicine, Guangzhou 510632, China |
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Abstract It is intending to prepare tumor necrosis factor α (TNFα) derivative TRSP10 which can lead prostate cancer DU145 cells apoptosis by genetic engineering and to study the inhibitory effect on DU145 cells in vitro. Trsp 10 gene sequences were synthesized by overlap extension PCR and then inserted into the site between SapI and NdeI in the highly-efficient expression vector,pKYB-MCS and conduct the optimal conditions to induce the expression of the fusion protein and set up the technology from vector construction to the expression and purification of the recombinant bacteria. MTT detect the inhibit proliferation effect of the TRSP10 on prostate cancer DU145 cells. The results showed that the optimal expression condition was as follows: at 0.8 mmol /L IPTG,37 ℃ and 8 hours.Puried by the IMPACT system and HPLC technology, the purity of the TRSP10 is 96%.The molecular mass is 3.59kDa that is determined by the mass spectrometry, which is consistent with the theoretical data. In vitro, studies showed that, TRSP10 could significantly inhibit the proliferation of prostate cancer DU145 cells.The rates of inhibition among the concentrations of 5,10,20,40μmol/L, are 11.40%, 22.97%, 33.26%, 48.35% respectively.
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Received: 04 January 2015
Published: 25 April 2015
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