Reference genes selection for quantitative real-time PCR using RankAggreg method in different tissues of Capra hircus.
Reference genes selection for quantitative real-time PCR using RankAggreg method in different tissues of Capra hircus.
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DOI：
10.1371/journal.pone.0083041
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发表时间：
2013
期刊：
影响因子：
3.7
通讯作者：
Bakhtiarizadeh MR
中科院分区：
文献类型：
--
作者：
Najafpanah MJ;Sadeghi M;Bakhtiarizadeh MR
鉴定具有稳定基因表达水平的内参基因是使用实时定量 PCR (RT-qPCR) 分析基因表达数据获得可靠结果的重要先决条件。由于参考基因的基本假设是在所有样本类型中以完全相同的水平表达，因此在本研究中，我们评估了九种最常用的内源对照（GAPDH、ACTB、18S rRNA、RPS18、HSP-90、ALAS、HMBS、ACAC 和 B2M）在家山羊 Capra hircus 的四种不同组织中的表达稳定性，包括肝脏、内脏、皮下脂肪和最长肌。治疗（使用 NRC 计算机软件准备的标准饮食作为对照，以及相同的饮食加 1 毫克铬/天）。我们使用六种不同的软件程序对参考基因进行排序，发现它们之间的基因的个体排序不同。此外，不同组织中所研究基因的排序模式存在显着差异。采用排名聚合方法将六个项目的排名列表合并成共识排名。我们的结果表明，HSP-90 几乎总是所有研究组织中两个最稳定的基因之一。因此，建议对山羊中的 RT-qPCR 数据进行准确标准化，而 GAPDH、ACTB 和 RPS18 显示出最多变化的表达，应避免作为内参基因。
Identification of reference genes with stable levels of gene expression is an important prerequisite for obtaining reliable results in analysis of gene expression data using quantitative real time PCR (RT-qPCR). Since the underlying assumption of reference genes is that expressed at the exact same level in all sample types, in this study, we evaluated the expression stability of nine most commonly used endogenous controls (GAPDH, ACTB, 18S rRNA, RPS18, HSP-90, ALAS, HMBS, ACAC, and B2M) in four different tissues of the domestic goat, Capra hircus, including liver, visceral, subcutaneous fat and longissimus muscles, across different experimental treatments (a standard diet prepared using the NRC computer software as control and the same diet plus one mg chromium/day). We used six different software programs for ranking of reference genes and found that individual rankings of the genes differed among them. Additionally, there was a significant difference in ranking patterns of the studied genes among different tissues. A rank aggregation method was applied to combine the ranking lists of the six programs to a consensus ranking. Our results revealed that HSP-90 was nearly always among the two most stable genes in all studied tissues. Therefore, it is recommended for accurate normalization of RT-qPCR data in goats, while GAPDH, ACTB, and RPS18 showed the most varied expressions and should be avoided as reference genes.
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影响因子：
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作者：
Becker, Christiane;Riedmaier, Irmgard;Meyer, Heinrich H. D.
通讯作者：
Meyer, Heinrich H. D.
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Martens, Ann
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通讯作者：
Bustin, SA
影响因子：
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Lallemant B;Evrard A;Combescure C;Chapuis H;Chambon G;Raynal C;Reynaud C;Sabra O;Joubert D;Hollande F;Lallemant JG;Lumbroso S;Brouillet JP
通讯作者：
Brouillet JP