Chen Yu-Chia, Kao Shang-Ching, Chou Hsiao-Chin, Lin Wei-Hsiang, Wong Feng-Hua, Chow Wei-Yuan
Department of Life Sciences, National Yang-Ming University, Shih-Pai, Taipei 112, Taiwan.
Anal Biochem. 2008 Apr 1;375(1):46-52. doi: 10.1016/j.ab.2007.12.037. Epub 2008 Jan 9.
In this study, a quantitative PCR (qPCR) method was developed to determine the A-to-I RNA editing frequencies at specific sites. The A-to-I RNA editing of nuclear transcripts exerts profound effects on the biological activities of gene products. RNA editing of nuclear gene transcripts have been shown to be developmentally regulated and tissue specific, and alternations of RNA editing activities have been observed under pathological conditions. Two sites of ionotropic glutamate receptor subunits, the Q/R site of zebrafish gria2alpha and the Y/C site of grik2alpha, were chosen in this study to demonstrate the applicability of the SYBR Green detection-based real-time PCR method to measure RNA editing activities during zebrafish development. The results obtained by qPCR were consistent with those obtained by the limited primer extension. However, the qPCR method has the advantages of easy handling and low cost.
在本研究中，开发了一种定量PCR（qPCR）方法来测定特定位点的A-to-I RNA编辑频率。核转录本的A-to-I RNA编辑对基因产物的生物学活性具有深远影响。已表明核基因转录本的RNA编辑受到发育调控且具有组织特异性，并且在病理条件下观察到RNA编辑活性的改变。本研究选择了离子型谷氨酸受体亚基的两个位点，即斑马鱼gria2alpha的Q/R位点和grik2alpha的Y/C位点，以证明基于SYBR Green检测的实时PCR方法在测量斑马鱼发育过程中RNA编辑活性方面的适用性。通过qPCR获得的结果与通过有限引物延伸获得的结果一致。然而，qPCR方法具有操作简便和成本低的优点。
