Screening of Genes Involved in Hepatoma Cells SMMC-7721 Stress Response Induced by Different Stressors
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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GeneAtlas基因芯片系统，制作Affymetrix Human Gene 1.1 ST Array Strip表达谱芯片，采用iterPlier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qRT-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；qRT-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qRT-PCR检测中变化趋势较一致的应激相关基因共有17个。
Abstract:ObjectiveTo 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.
MethodsWe 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 72h and 4h treatment, we collected the cells and extracted total RNA. The GeneAtlas microarray system was used to make Affymetrix Human Gene 1.1 ST Array Strip genechip. iterPlier 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, qRT-PCR was used to detect the relative expression of each gene.
ResultsEventually 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 qRT-PCR.
ConclusionIn 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 qRT-PCR.
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Key words:
- Hepatocellular carcinoma cells /
- Stress-related genes /
- Genechip /
- qRT-PCR
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表 1 芯片读数计算值一致上调的18个基因
Table 1 18 genes with consistently up-regulated expression in genechip readings calculated value
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表 2 芯片读数计算值一致下调的14个基因
Table 2 14 genes with consistently down-regulated expression in genechip readings calculated value
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表 3 芯片读数计算值互有上下调的16个基因
Table 3 16 genes with inconsistently regulated expression in genechip readings calculated value
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表 4 芯片读数计算值无显著变化的5个基因
Table 4 Five genes without significantly changed expression in genechip readings calculated value
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