Expression of recombinant human soluble apoptosis 2 ligand and vitro experimental study on laryngeal carcinoma cell line Hepa2

OBJECTIVE:To investigate the specificity role of the apoptosis factor TRAIL gene with telomerase promoter in tumor gene therapy. METHODS:PBLs(Peripheral Blood lymphocyte) were stimulated for proliferation and the total RNA was extracted. shTRAIL (recombinant humam soluble APO2),TRAIL gene with interleukin 2 signal peptide, was amplified by RT-PCR and cloned into the vector pGL3-181h TERT promoter downstream to form an eukaryotic expressing vector. Expressing protein was identified by Western blot on laryngeal carcinoma cell line Hepa2. The morphological changes were observed by transmission electron microscope (TEM). The apoptotic rates and the changes of cell cycle were measured by flow cytometry (FCM) in cell line Hepa2 and HL-7702. RESULTS: The sequence of 613 bp cDNA was obtained and identified by enzyme digestion and sequencing analysis, and the recombinant eukaryotic expression vector for TRAIL gene was successfully constructed. The results of the Western blot showed the transfected Hepa2 cells expressed TRAIL protein and apoptosis were induced. CONCLUSION:Successful construction of eukaryotic expression vector pGL3-181hTERT/TRAIL provides the possibility for gene therapy of tumor.