Miraglia Loren J, King Frederick J, Damoiseaux Robert
Department of Genomics, The Genomics Institute of the Novartis Research Foundation, San Diego, California, USA.
Comb Chem High Throughput Screen. 2011 Sep;14(8):648-57. doi: 10.2174/138620711796504389.
The luminescent reporter gene assay (LRGA) is arguably the most prominent type of reporter gene assay used in biomolecular and pharmaceutical development laboratories. Part of this popularity is due to the high signal associated with luciferases, the foundation of this technology. This feature makes them ideally suited for high throughput screening applications where potentially millions of chemical compounds can be analyzed in a given assay. Recent technical advancements that enhance signal stability of the luciferases along with development and commercialization of multiple forms of luciferases, their respective substrates, and improvements in expression vectors for reporter gene assay (RGA) applications have broadened their use. While the practical challenges related to the application of luminescent technology in a laboratory setting have been overcome, there remains much to do in laying a systematic approach towards the construction of RGAs, which are essential to the elucidation of the basic biology for genes of interest. This mini-review aims at giving a birds-eye view of the available luciferases, substrates and other luminescent technologies available and provides a general blueprint as well as practical considerations for constructing and interfacing RGAs with chemical biology and functional genomics for the elucidation of fundamental biological questions and for biomedical research.
发光报告基因检测（LRGA）可以说是生物分子和药物开发实验室中使用的最突出的报告基因检测类型。其受欢迎的部分原因在于与荧光素酶相关的高信号，这是该技术的基础。这一特性使其非常适合高通量筛选应用，在给定的检测中可以分析潜在的数百万种化合物。最近的技术进步提高了荧光素酶的信号稳定性，同时多种形式的荧光素酶、它们各自的底物得以开发和商业化，以及报告基因检测（RGA）应用的表达载体得到改进，这些都扩大了它们的用途。虽然在实验室环境中应用发光技术的实际挑战已经克服，但在建立一种系统的方法来构建RGA方面仍有许多工作要做，而RGA对于阐明感兴趣基因的基础生物学至关重要。本综述旨在鸟瞰现有的荧光素酶、底物和其他发光技术，并提供一个总体蓝图以及实际考虑因素，用于构建RGA并将其与化学生物学和功能基因组学相结合，以阐明基本生物学问题和进行生物医学研究。
