Wang Dan Ohtan, Okamoto Akimitsu
Institute for Integrated Cell-Material Sciences (WPI-iCeMS), Kyoto University, Yoshida-Honmachi, Sakyo-ku, Kyoto, 606-8501, Japan,
Methods Mol Biol. 2015;1262:69-87. doi: 10.1007/978-1-4939-2253-6_5.
Engineered probes to adapt new photochemical properties upon recognition of target nucleic acids offer powerful tools to DNA and RNA visualization technologies. Herein, we describe a rapid and effective visualization method of nucleic acids in both fixed and living cells with hybridization-sensitive fluorescent oligonucleotide probes. These probes are efficiently quenched in an aqueous environment due to the homodimeric, excitonic interactions between fluorophores but become highly fluorescent upon hybridization to DNA or RNA with complementary sequences. The fast hybridization kinetics and quick fluorescence activation of the new probes allow applications to simplify the conventional fluorescent in situ hybridization protocols and reduce the amount of time to process the samples. Furthermore, hybridization-sensitive fluorescence emission of the probes allows monitoring dynamic behaviors of RNA in living cells.
经设计的探针在识别靶核酸后能适应新的光化学性质,为DNA和RNA可视化技术提供了强大工具。在此,我们描述了一种利用杂交敏感荧光寡核苷酸探针在固定细胞和活细胞中快速有效地可视化核酸的方法。由于荧光团之间的同二聚体激子相互作用,这些探针在水性环境中被有效淬灭,但与具有互补序列的DNA或RNA杂交后会变得高度荧光。新探针快速的杂交动力学和快速的荧光激活特性使得其可用于简化传统的荧光原位杂交方案,并减少样品处理时间。此外,探针的杂交敏感荧光发射能够监测活细胞中RNA的动态行为。