SHEN Qi,ZHANG Wenwen,TAO Xuejiao, et al. Construction of a 14-3-3γ expression vector and its effect on proliferation and apoptosis of uterine leiomyoma cells[J]. JOURNAL OF WEZHOU MEDICAL UNIVERSITY, 2014, 44(2): 79-.
Abstract：Objective: To construct stably and highly expressing 14-3-3γgene eukaryotic expression plasmid vector and to explore its antitumor effects on uterine leiomyoma cells in vitro. Methods: The total RNA was isolated from the uterine leiomyoma cells. After reverse transcription reaction, 14-3-3γwas amplified by RT-PCR. The PCR product was cloned into the eukaryote expression vector pCMV-N-Flag, the resulted recombinant was designated as 14-3-3γ-pCMV-N-Flag. After sequence determination, the recombinant vector was transfected into uterine leiomyoma cells by lipofectin. The expression of the interesting protein was confirmed by Western Blot analysis. Cell proliferation was assessed by cell counting kit-8 assay and the percentage of apoptotic cells detected by annexinV-FITC/PI double-staining was determined by flow cytometry. Results: ①The 14-3-3γgene eukaryotic expression vector was successfully constructed and identified by sequencing. ②The expression level of 14-3-3γprotein was increased obviously after transfection (P<0.05). ③The inhibitory effect on the rate of cell proliferation was 52.90% after transfection (P<0.05). ④After transfection, the percentage of apoptotic cells was increased obviously (P<0.05). Conclusion: The 14-3-3γ-pCMV-N-Flag gene eukaryotic expression plasmid vector is successfully constructed which could suppress proliferation and induce apoptosis of uterine leiomyoma cells.
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