National Key Facility for Crop Gene Resources and Genetic Improvement, ICS, CAAS, 12 Zhongguancun South Street, Haidian District, Beijing, 100081, China.
Anal Biochem. 2021 Jan 1;612:114021. doi: 10.1016/j.ab.2020.114021. Epub 2020 Nov 12.
We developed a binary vector co-expressing firefly luciferase (FF) and Renilla luciferase (REN) to detect protein stability in response to different stimuli, and verified the functionality of the vector. The StrigoQuant-like reporter expressing FF and REN in one transcript is a sensitive tool for detecting protein abundance in different genotypes. However, we found that significant differences in the relative FF/REN ratio of empty StrigoQuant vector in different genotypes. Therefore, to determine the actual protein abundance, the relative FF/REN ratio of the protein of interest should be normalized to that of the empty vector.
我们开发了一种共表达萤火虫荧光素酶 (FF) 和海肾荧光素酶 (REN) 的二元载体，以检测对不同刺激的蛋白质稳定性，并验证了该载体的功能。在一个转录本中表达 FF 和 REN 的 StrigoQuant 样报告基因是检测不同基因型中蛋白质丰度的敏感工具。然而，我们发现不同基因型中空的 StrigoQuant 载体的相对 FF/REN 比值存在显著差异。因此，为了确定实际的蛋白质丰度，应该将感兴趣的蛋白质的相对 FF/REN 比值与空载体的比值进行归一化。
