摘要
目的筛选体外培养的肝癌SMMC-7721细胞在抵御不同应激原而恶性增殖的过程中,与应激反应密切相关的基因。方法常规体外培养肝癌SMMC-7721细胞,将处于对数生长期的细胞分为正常组、G418组和氯化钙组,除正常组外,其余两组分别采用G418、氯化钙加以干预,于72和4 h后收集各组肝癌细胞,抽提总RNA,使用Gene Atlas基因芯片系统,制作Affymetrix Human Gene 1.1ST Array Strip表达谱芯片,采用iter Plier法计算基因表达读数计算值,并进行统计学处理,观察与肝癌细胞应激反应相关的基因群的差异性表达特征,通过q RT-PCR检测各基因的相对表达量,对芯片结果加以验证。结果芯片检测筛选到53个表达量大、且在G418和氯化钙作用下表达量变化较一致的基因;q RT-PCR验证的53个应激相关基因中,17个基因与基因芯片结果完全吻合。结论在肝癌SMMC-7721细胞抵御G418和氯化钙应激的过程中,作用较关键且在基因芯片和q RT-PCR检测中变化趋势较一致的应激相关基因共有17个。
Objective To screen the genes which are related to stress response closely in the malignant proliferation of hepatocellular carcinoma cells SMMC-7721 against different stressors in vitro. Methods We conventionally cultured SMMC-7721 cells in vitro, grouping the cells in log phase into normal group, G418 group and calcium group. Each treatment group was given the corresponding drug respectively. After 72 h and 4h treatment, we collected the cells and extracted total RNA. The Gene Atlas microarray system was used to make Affymetrix Human Gene 1.1 ST Array Strip genechip. iter Plier was used for calculating gene expression, to work out preliminary statistical processing. We observed the characteristics of genes expression related to stress response in hepatocellular carcinoma cells. Then, q RT-PCR was used to detect the relative expression of each gene. Results Eventually we screened 53 genes which had a large quantity of expression and the change were consistent with those under the effect of G418 and calcium chloride, moreover, 17 genes had relatively consistent trend with genechips verified by q RT-PCR. Conclusion In the process of hepatocellular carcinoma cells SMMC-7721 resisting G418 and calcium chloride stress, 17 genes are important and verified relatively consistent trend by genechips and q RT-PCR.
出处
《肿瘤防治研究》
CAS
CSCD
北大核心
2016年第6期463-467,共5页
Cancer Research on Prevention and Treatment
基金
国家自然科学基金(81273641)
中国博士后科学基金(2013M531201)
上海市科委博士后科研基金项目(13R21415900)
上海高校青年教师培养资助计划(ZZszy13004)
关键词
肝癌细胞
应激基因
基因芯片
QRT-PCR
Hepatocellular carcinoma cells
Stress-related genes
Genechip
qRT-PCR
