Sando Shinsuke, Kool Eric T
Department of Synthetic Chemistry and Biological Chemistry, Faculty of Engineering, Kyoto University, Kyoto 606-8501, Japan.
Nucleic Acids Res Suppl. 2002(2):121-2. doi: 10.1093/nass/2.1.121.
We report on a new fluorescence reporting strategy in which dabsyl, a well-known quencher, activates a hydroxyl group in a probe to convert it to a leaving group. When a nucleophilic phosphorothioate probe binds adjacent to a dabsyl quenched probe, autoligation occurs, releasing the quencher, and lighting up the probes. These self-ligating DNA probes were used for sequence-specific detection of 16S rRNA in E. coli cells. Strong fluorescence was observed only when the phosphorothioate and quenched dabsyl probes bind side-by-side on a 16S rRNA target. The results demonstrate the use of QUAL probes to detect specific RNA sequences in bacterial cells without enzymes and without washing steps.
我们报道了一种新的荧光报告策略，其中，一种著名的淬灭剂达磺酰激活探针中的一个羟基，将其转化为离去基团。当亲核硫代磷酸酯探针与达磺酰淬灭探针相邻结合时，会发生自动连接，释放淬灭剂，并使探针发光。这些自连接DNA探针用于大肠杆菌细胞中16S rRNA的序列特异性检测。只有当硫代磷酸酯探针和淬灭的达磺酰探针在16S rRNA靶标上并排结合时，才观察到强荧光。结果表明，使用QUAL探针可在无酶且无需洗涤步骤的情况下检测细菌细胞中的特定RNA序列。
