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目的构建MASPIN真核表达载体,分析MASPIN过表达对胃腺癌细胞株SGC7901的影响。方法PCR扩增MASPIN基因全长,用载体PCR2.1构建重组质粒,转染至胃癌细胞株SGC7901。RT-PCR、Western blot检测MASPIN基因表达变化,MTT法检测细胞增殖情况,流式细胞仪(FCM)分析细胞周期,观察过表达MASPIN对SGC7901细胞株的影响。结果成功构建MASPIN/PCR2.1表达载体并转染至胃癌细胞株SGC7901,转染上调了SGC7901细胞株MASPIN在mRNA、蛋白水平的表达。转染MASPIN的SGC7901细胞增殖率随时间而减慢,第24、48、72小时的细胞增殖率分别为93.07%、81.97%、66.5%,明显低于转染空载体组(95.98%、95.79%、95.59%,P<0.05)。与转染空载体组相比,过表达MASPIN的SGC7901细胞株在G0/G1期所占比例明显提高(70.3%vs55.6%),S期细胞减少(19.8%vs34.9%,P<0.05)。结论成功构建MASPIN/PCR2.1表达载体,过表达MASPIN可抑制SGC7901细胞株的增殖。
Objective To construct MASPIN eukaryotic expression vector and analyze the effect of MASPIN overexpression on gastric adenocarcinoma cell line SGC7901. Methods The full-length MASPIN gene was amplified by PCR. The recombinant plasmid was constructed by PCR2.1 and transfected into gastric cancer cell line SGC7901. The expression of MASPIN gene was detected by RT-PCR and Western blot. The cell proliferation was detected by MTT assay. The cell cycle was analyzed by flow cytometry (FCM). The effect of over-expressed MASPIN on SGC7901 cell line was observed. Results The MASPIN / PCR2.1 expression vector was successfully constructed and transfected into gastric cancer cell line SGC7901. Transfection enhanced the expression of MASPIN mRNA and protein in SGC7901 cell line. The proliferation rate of SGC7901 cells transfected with MASPIN slowed down with time, and the cell proliferation rates at the 24th, 48th and 72th hour were 93.07%, 81.97% and 66.5%, respectively, which were significantly lower than that of the transfected empty vector group (95.98%, 95.79% , 95.59%, P <0.05). The percentage of SGC7901 cells overexpressing MASPIN in G0 / G1 phase increased significantly (70.3% vs 55.6%) and S phase cells decreased (19.8% vs 34.9%, P <0.05) ). Conclusion MASPIN / PCR2.1 expression vector was constructed successfully. Over-expression of MASPIN inhibited the proliferation of SGC7901 cells.