LAGENBIO-I3A, Instituto Aragonés de Ciencias de Salud, University of Zaragoza, Miguel Servet 177, 50013 Zaragoza, Spain.
Biochem Biophys Res Commun. 2011 Apr 22;407(4):758-63. doi: 10.1016/j.bbrc.2011.03.096. Epub 2011 Mar 31.
Reliability and accuracy of real-time quantitative PCR results depend on the use of housekeeping genes which must be constitutively expressed thorough the samples of the study. In the present work, we tested the expression stability of six candidate housekeeping genes (Actb, Rn18s, Gapdh, Hprt1, Sdha and B2m) considering sex, age, muscle-type and neurodegeneration or denervation status in mouse muscle satellite cells. Their expression varied under all variables tested; therefore the ranking of the most suitable genes for the normalization is modified depending on the factors included in the analysis, especially the age of the donor. Moreover, we describe the unsuitability of Rn18s in analysis comprising samples of different ages. On the other hand, we demonstrate that the use of the two best genes in each case is enough to obtain a reliable normalization factor. In this work, we give a broad information of the best housekeeping genes in mouse myogenic cells depending on the variables included in the experimental design.
实时定量 PCR 结果的可靠性和准确性取决于管家基因的使用,这些基因必须在研究样本中持续表达。在本工作中,我们测试了六个候选管家基因(Actb、Rn18s、Gapdh、Hprt1、Sdha 和 B2m)在考虑性别、年龄、肌肉类型以及神经退行性变或去神经状态的情况下在小鼠肌肉卫星细胞中的表达稳定性。它们的表达在所有测试的变量下都有所不同;因此,根据分析中包含的因素(特别是供体的年龄),对最适合基因进行归一化的排序进行了修改。此外,我们还描述了 Rn18s 在包含不同年龄样本的分析中的不适用性。另一方面,我们证明在每种情况下使用两个最佳基因足以获得可靠的归一化因子。在这项工作中,我们根据实验设计中包含的变量,为小鼠成肌细胞中的最佳管家基因提供了广泛的信息。