Validation of endogenous reference genes for qRT-PCR analysis of human visceral adipose samples.
Validation of endogenous reference genes for qRT-PCR analysis of human visceral adipose samples.
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DOI：
10.1186/1471-2199-11-39
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发表时间：
2010-05-21
影响因子：
--
通讯作者：
Baranova A
中科院分区：
文献类型：
--
作者：
Mehta R;Birerdinc A;Hossain N;Afendy A;Chandhoke V;Younossi Z;Baranova A
鉴于肥胖在全球范围内的流行比例和代谢综合征的同时流行，迫切需要更好地理解代谢综合征的潜在机制，特别是可能参与肥胖、胰岛素抵抗和相关的一系列慢性肝病的基因表达差异。实时PCR（qRT-PCR）是研究不同组织和实验条件下相关基因表达变化的标准方法。然而，起始材料的量、酶促效率和抑制剂的存在的变化可导致定量误差。因此，准确的数据标准化至关重要。在几种已知的数据标准化策略中，使用参考基因作为内部对照是最常见的方法。最近的研究表明，肥胖和胰岛素抵抗的存在都会影响网膜脂肪中常用参考基因的表达。在这项研究中，我们验证了候选参考基因适用于qRT-PCR分析实验，使用内脏脂肪样本从肥胖和瘦的个人。使用三种流行的算法GeNorm、Normalization和BestKeeper交叉验证八个选定的参考基因的表达稳定性，发现ACTB和RPII是最稳定的参考基因。我们推荐ACTB和RPII作为最适合人类内脏脂肪组织基因表达研究的稳定参考基因。使用这些基因作为参考对可以进一步增强该模型系统中qRT-PCR的稳健性。
Given the epidemic proportions of obesity worldwide and the concurrent prevalence of metabolic syndrome, there is an urgent need for better understanding the underlying mechanisms of metabolic syndrome, in particular, the gene expression differences which may participate in obesity, insulin resistance and the associated series of chronic liver conditions. Real-time PCR (qRT-PCR) is the standard method for studying changes in relative gene expression in different tissues and experimental conditions. However, variations in amount of starting material, enzymatic efficiency and presence of inhibitors can lead to quantification errors. Hence the need for accurate data normalization is vital. Among several known strategies for data normalization, the use of reference genes as an internal control is the most common approach. Recent studies have shown that both obesity and presence of insulin resistance influence an expression of commonly used reference genes in omental fat. In this study we validated candidate reference genes suitable for qRT-PCR profiling experiments using visceral adipose samples from obese and lean individuals. Cross-validation of expression stability of eight selected reference genes using three popular algorithms, GeNorm, NormFinder and BestKeeper found ACTB and RPII as most stable reference genes. We recommend ACTB and RPII as stable reference genes most suitable for gene expression studies of human visceral adipose tissue. The use of these genes as a reference pair may further enhance the robustness of qRT-PCR in this model system.
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影响因子：
3.7
作者：
de Jonge HJ;Fehrmann RS;de Bont ES;Hofstra RM;Gerbens F;Kamps WA;de Vries EG;van der Zee AG;te Meerman GJ;ter Elst A
通讯作者：
ter Elst A
影响因子：
6.5
作者：
Linder, K;Arner, P;Norstedt, G
通讯作者：
Norstedt, G
影响因子：
4.1
作者：
Thellin, O;Zorzi, W;Heinen, E
通讯作者：
Heinen, E
影响因子：
3.5
作者：
Baranova, A;Hammarsund, M;Yankovsky, N
通讯作者：
Yankovsky, N
DOI：
10.1046/j.1467-789x.2000.00008.x
发表时间：
2000-05-01
期刊：
Obesity reviews : an official journal of the International Association for the Study of Obesity
影响因子：
--
作者：
Bosello, O;Zamboni, M
通讯作者：
Zamboni, M