YKL-40基因真核表达载体的构建及对人胃癌SGC-7901细胞增殖作用的影响  

Construction of eukaryotic expression vector of YKL-40 and its effects on proliferation of gastric cancer cell line SGC-7901

作　　者：庞春艳[1] 车团结[2] 李宁[3] 闫琨[1] 李琳[2] 王勤[3] 陈卫[1] 

机构地区：[1]兰州大学基础医学院生物化学与分子生物学研究所,甘肃兰州730000 [2]兰州大学生命科学院细胞生物学研究所,甘肃兰州730000 [3]兰州大学生命科学院生物物理研究所,甘肃兰州730000

出　　处：《第四军医大学学报》2008年第12期1121-1124,共4页Journal of the Fourth Military Medical University

摘　　要：目的:构建针对人类软骨糖蛋白-39(HCGP-39,YKL-40)的真核表达载体,观察其对肿瘤细胞增殖的影响.方法:采用逆转录方法从乳腺癌组织中获得基因YKL-40,双酶切后插入线性化的pcDNA3.1载体真核启动子下游,重组体经限制性内切酶及测序鉴定,将阳性克隆的载体转染胃癌细胞株SGC-7901,采用噻唑蓝(MTT)法检测细胞的增殖活性、细胞计数并观察细胞生长情况、半定量RT-PCR检测基因YKL-40 mRNA的表达、流式细胞术检测SGC-7901细胞的周期变化情况.结果:限制性内切酶及测序鉴定表明成功地构建了针对基因YKL-40的真核表达载体;将其转染胃癌细胞SGC-7901后,发现细胞增殖加快,差异具有统计学意义(P<0.05);RT-PCR检测基因YKL-40 mRNA的表达增加;流式细胞检测S期细胞增多.结论:成功地构建了针对基因YKL-40的真核表达载体,该载体可使胃癌细胞株SGC-7901增殖加快.AIM： To construct eukaryotic expression vector against human cartilage glycoprotein-39（ HCGP-39, YKL40） and to study its effect on proliferation of gastric cancer cell line SGC- 7901. METHODS： YKL40 was obtained from breast cancer tissue by RT-PCR method. The digested double strands DNA were inserted into the downstream of promoter of linearized pcDNA3.1. After it was confirmed by restrictive enzyme digestion method and DNA sequence analysis, the recombinant plasmid was transfected into SGC-7901, a gastric cancer cell line, by using calcium phosphate. The cell proliferation was determined by MTT method. The level of YKL40 mRNA in SGC-7901 cells was detected by RT- PCR assay. Cell cycles were assayed by FCM analysis. RESULTS： The expression vector of YKL-40 was constructed successfully. The recombinant plasmid enhanced the proliferation of SGC-7901 cells dramatically. The result of RT-PCR showed that YKL40 mRNA level was increased evidently. The percentage of cells in S phase increased. CONCLUSION： The eukaryotie expression vector of YKL40 is successfully constructed and it can enhance the proliferation of SGC-7901 cells.

关 键 词：YKL-40 HCGP-39 胃肿瘤 

分 类 号：R73[医药卫生—肿瘤]