The fluorescence reaction based on the oligonucleotide (ON) template reaction is one of the most basic working principles of the RNA detection probe. In this method, an ON probe having a reactive functional group and an ON probe having a fluorescent precursor whose fluorescence is suppressed by a protecting group are associated on the target RNA, and then the protective group is removed and fluorescence signal is generated. Since this fluorescence reaction specifically occurs in the presence of target RNA, it can be the working principle RNA detection probe. In order to detect intracellular RNA highly sensitively, we worked on (1) accelerating ON template reaction and (2) development of a new method of intracellular administration of ON probe without lipofection. Prospective results were obtained in (1) by appropriately selecting the nucleophile for the reaction, and in (2) by use of phosphorothioate in the ON probe.