Tang Rongying, Dodd Andrew, Lai Daniel, McNabb Warren C, Love Donald R
School of Biological Sciences, University of Auckland, Private Bag 92019, Auckland 1142, New Zealand.
Acta Biochim Biophys Sin (Shanghai). 2007 May;39(5):384-90. doi: 10.1111/j.1745-7270.2007.00283.x.
The normalization of quantitative real time RT-PCR (qRT-PCR) is important to obtain accurate gene expression data. The most common method for qRT-PCR normalization is to use reference, or housekeeping genes. However, there is emerging evidence that even reference genes can be regulated under different conditions. qRT-PCR has only recently been used in terms of zebrafish gene expression studies and there is no validated set of reference genes. This study characterizes the expression of nine possible reference genes during zebrafish embryonic development and in a zebrafish tissue panel. All nine reference genes exhibited variable expression. The beta-actin, EF1alpha and Rpl13alpha genes comprise a validated reference gene panel for zebrafish developmental time course studies, and the EF1alpha, Rpl13alpha and 18S rRNA genes are more suitable as a reference gene panel for zebrafish tissue analysis. Importantly, the zebrafish GAPDH gene appears unsuitable as reference gene for both types of studies.
定量实时逆转录聚合酶链反应（qRT-PCR）的标准化对于获得准确的基因表达数据很重要。qRT-PCR标准化最常用的方法是使用内参基因或管家基因。然而，越来越多的证据表明，即使是内参基因在不同条件下也可能受到调控。qRT-PCR直到最近才用于斑马鱼基因表达研究，并且还没有经过验证的内参基因集。本研究描述了九个可能的内参基因在斑马鱼胚胎发育过程中和斑马鱼组织样本中的表达情况。所有九个内参基因均表现出可变表达。β-肌动蛋白、EF1α和Rpl13α基因构成了用于斑马鱼发育时间进程研究的经过验证的内参基因组合，而EF1α、Rpl13α和18S rRNA基因更适合作为斑马鱼组织分析的内参基因组合。重要的是，斑马鱼GAPDH基因似乎不适用于这两种类型的研究作为内参基因。
