Ullmannová V, Haskovec C
Department of Molecular Genetics, Institute of Haematology and Blood Transfusion, Prague, Czech Republic.
Folia Biol (Praha). 2003;49(6):211-6.
Quantitative real-time RT-PCR is a very useful technique for estimating gene expression at the mRNA level. The expression of a tested gene has to be compared with that of a control gene. Various housekeeping genes have been used as control genes in different systems. In our study we tested several housekeeping genes in the model of gene expression after induction of apoptosis and differentiation. The myeloid cell lines were incubated with phorbol esters, butyric acid and combination of TNFalpha and IFNgamma to induce differentiation. Camptothecin was used for induction of apoptosis. Tested control genes included beta2-microglobulin, GAPDH, 18S ribosomal RNA and abl. GAPDH was found to be the best control gene in the apoptotic system. Different control genes were suitable for different systems where differentiation or senescence was induced. Our results show that attention should be paid to the choice of an appropriate control gene of quantitative real-time RT-PCR for different experimental models and various experimental conditions.
定量实时逆转录聚合酶链反应（Quantitative real-time RT-PCR）是一种在mRNA水平估计基因表达的非常有用的技术。测试基因的表达必须与对照基因的表达进行比较。在不同系统中，各种管家基因已被用作对照基因。在我们的研究中，我们在诱导凋亡和分化后的基因表达模型中测试了几个管家基因。髓样细胞系与佛波酯、丁酸以及肿瘤坏死因子α（TNFalpha）和干扰素γ（IFNgamma）的组合一起孵育以诱导分化。喜树碱用于诱导凋亡。测试的对照基因包括β2-微球蛋白、甘油醛-3-磷酸脱氢酶（GAPDH）、18S核糖体RNA和abl。发现GAPDH是凋亡系统中最佳的对照基因。不同的对照基因适用于诱导分化或衰老的不同系统。我们的结果表明，对于不同的实验模型和各种实验条件，应注意选择定量实时RT-PCR合适的对照基因。
