Blizard Institute of Cell and Molecular Science, Barts and the London School of Medicine and Dentistry, Queen Mary University of London, Whitechapel, London E1 1BB, UK.
Methods. 2010 Apr;50(4):217-26. doi: 10.1016/j.ymeth.2009.12.006. Epub 2009 Dec 16.
The polymerase chain reaction (PCR) has matured from a labour- and time-intensive, low throughput qualitative gel-based technique to an easily automated, rapid, high throughput quantitative technology. Real-time quantitative PCR (qPCR) has become the benchmark technology for the detection and quantification of nucleic acids in a research, diagnostic, forensic and biotechnology setting. However, ill-assorted pre-assay conditions, poor assay design and inappropriate data analysis methodologies have resulted in the recurrent publication of data that are at best inconsistent and at worst irrelevant and even misleading. Furthermore, there is a lamentable lack of transparency of reporting, with the "Materials and Methods" sections of many publications, especially those with high impact factors, not fit for the purpose of evaluating the quality of any reported qPCR data. This poses a challenge to the integrity of the scientific literature, with serious consequences not just for basic research, but potentially calamitous implications for drug development and disease monitoring. These issues are being addressed by a set of guidelines that propose a minimum standard for the provision of information for qPCR experiments ("MIQE"). MIQE aims to restructure to-day's free-for-all qPCR methods into a more consistent format that will encourage detailed auditing of experimental detail, data analysis and reporting principles. General implementation of these guidelines is an important requisite for the maturing of qPCR into a robust, accurate and reliable nucleic acid quantification technology.
聚合酶链反应 (PCR) 已经从一种劳动强度大、时间长、高通量基于凝胶的定性技术发展成为一种易于自动化、快速、高通量的定量技术。实时定量 PCR(qPCR) 已成为研究、诊断、法医和生物技术领域检测和定量核酸的基准技术。然而，不适当的预实验条件、不良的实验设计和不适当的数据分析方法导致反复发表的数据要么不一致，要么最差是不相关甚至误导。此外，报告的透明度很差，许多出版物，尤其是那些具有高影响因子的出版物，“材料和方法”部分不适合评估任何报告的 qPCR 数据的质量。这对科学文献的完整性提出了挑战，不仅对基础研究产生严重影响，而且对药物开发和疾病监测也可能产生灾难性影响。一组指南正在解决这些问题，这些指南为 qPCR 实验提出了最低信息要求 (MIQE)。MIQE 的目的是将当今的 qPCR 方法进行重构，形成更一致的格式，以鼓励对实验细节、数据分析和报告原则进行详细审查。这些指南的普遍实施是将 qPCR 成熟为一种强大、准确和可靠的核酸定量技术的重要前提。
