Haugwitz Michael, Nourzaie Omar, Garachtchenko Tatiana, Hu Lanrong, Gandlur Suvarna, Olsen Cathy, Farmer Andrew, Chaga Grigoriy, Sagawa Hiroaki
Clontech Laboratories, Inc., 1290 Terra Bella Ave., Mountain View, CA 94043, USA.
Curr Chem Genomics. 2008 Feb 25;1:11-9. doi: 10.2174/1875397300801010011.
Reporter proteins are valuable tools to monitor promoter activities and characterize signal transduction pathways. Many of the currently available promoter reporters have drawbacks that compromise their performance. Enzyme-based reporter systems using cytosolic luciferases are highly sensitive, but require a cell lysis step that prevents their use in long-term monitoring. By contrast, secreted bioluminescent reporters like Metridia luciferase and Secreted Alkaline Phosphatase can be assayed repeatedly, using supernatant from the same live cell population to produce many sets of data over time. This is crucial for studies with limited amounts of cells, as in the case of stem cells. The use of secreted bioluminescent reporters also enables broader applications to provide more detailed information using live cells; for example, multiplexing with fluorescent proteins. Here, data is presented describing the characteristics of secreted Metridia luciferase and its use in multiplexing applications with either Secreted Alkaline Phosphatase or a fluorescent protein.
报告蛋白是监测启动子活性和表征信号转导途径的重要工具。目前许多可用的启动子报告基因都存在一些缺点，影响了它们的性能。基于酶的报告系统使用胞质荧光素酶，灵敏度很高，但需要进行细胞裂解步骤，这使得它们无法用于长期监测。相比之下，分泌型生物发光报告基因，如北极磷光素酶和分泌型碱性磷酸酶，可以反复检测，使用来自同一活细胞群体的上清液，随着时间的推移产生多组数据。这对于细胞数量有限的研究至关重要，例如干细胞研究。使用分泌型生物发光报告基因还可以实现更广泛的应用，利用活细胞提供更详细的信息；例如，与荧光蛋白进行多重检测。在此，本文展示了有关分泌型北极磷光素酶的特性及其在与分泌型碱性磷酸酶或荧光蛋白的多重检测应用中的数据。
