Bruckert G, Vivien D, Docagne F, Roussel B D
INSERM, INSERM U919, Serine Proteases and Pathophysiology of the Neurovascular Unit, GIP CYCERON, University Caen Basse Normandie, boulevard Becquerel, 14074, Caen, France.
Mol Neurobiol. 2016 Apr;53(3):1540-1550. doi: 10.1007/s12035-015-9114-5. Epub 2015 Feb 7.
Reverse transcription quantitative-polymerase chain reaction (RT-qPCR) has become a routine method in many laboratories. Normalization of data from experimental conditions is critical for data processing and is usually achieved by the use of a single reference gene. Nevertheless, as pointed by the Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines, several reference genes should be used for reliable normalization. Ageing is a physiological process that results in a decline of many expressed genes. Reliable normalization of RT-qPCR data becomes crucial when studying ageing. Here, we propose a RT-qPCR study from four mouse brain regions (cortex, hippocampus, striatum and cerebellum) at different ages (from 8 weeks to 22 months) in which we studied the expression of nine commonly used reference genes. With the use of two different algorithms, we found that all brain structures need at least two genes for a good normalization step. We propose specific pairs of gene for efficient data normalization in the four brain regions studied. These results underline the importance of reliable reference genes for specific brain regions in ageing.
逆转录定量聚合酶链反应（RT-qPCR）已成为许多实验室的常规方法。对来自实验条件的数据进行标准化处理对于数据处理至关重要，通常通过使用单个参照基因来实现。然而，正如《实时定量PCR实验发表的最低信息》（MIQE）指南所指出的，为了进行可靠的标准化，应该使用多个参照基因。衰老作为一种生理过程，会导致许多表达基因的减少。在研究衰老时，对RT-qPCR数据进行可靠的标准化处理变得至关重要。在此，我们提出一项针对不同年龄（8周龄至22月龄）的小鼠四个脑区（皮质、海马体、纹状体和小脑）的RT-qPCR研究，其中我们研究了九个常用参照基因的表达。通过使用两种不同的算法，我们发现所有脑结构至少需要两个基因才能实现良好的标准化步骤。我们针对所研究的四个脑区提出了用于有效数据标准化的特定基因对。这些结果强调了在衰老过程中针对特定脑区使用可靠参照基因的重要性。
