Characterization of the Triplet State of Hybridization-Sensitive DNA Probe by Using Fluorescence Correlation Spectroscopy
Characterization of the Triplet State of Hybridization-Sensitive DNA Probe by Using Fluorescence Correlation Spectroscopy
复制标题
使用荧光相关光谱表征杂交敏感 DNA 探针的三重态
DOI：
10.1021/jp307018k
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发表时间：
2013
期刊：
影响因子：
2.9
通讯作者：
C. -G.
中科院分区：
文献类型：
--
作者：
Shin;H. -S. ; Okamoto;A. ; Sako;Y. ; Kim;S. W. ; Kim;S. Y. ; Pack;C. -G.
新合成的杂交敏感DNA探针的非辐射弛豫机制至今尚未完全了解。在这项研究中，瞬态暗态的探针，这是一个双荧光染料连接到一个特定的DNA序列，使用荧光相关光谱（FCS）进行了研究。在已知影响DNA探针的系间交叉或光异构化的各种溶剂中测量瞬态暗态。在实验结果的基础上，建立了一个简化的探针的两能态模型，该模型为深入研究荧光团的非辐射弛豫机制及其在DNA和RNA检测中的应用提供了理论依据.在几微秒的时间尺度内测量的瞬态暗态是三重态，并且与光异构化、粘度、氧浓度和杂交有关，所有这些都是细胞显微镜的重要参数。在亚微秒的时间尺度中的瞬态暗态在添加靶DNA之后敏感地改变。该表征可以通过使用FCS提高探针识别靶DNA/RNA的能力，因为杂交后发生的三重态在时间尺度上与杂交前发生的三重态不同。
The nonradiative relaxation mechanism of the newly synthesized hybridized-sensitive DNA probe has not been fully understood until now. In this study, the transient dark state of the probe, which is a double fluorescent dye attached to a specific DNA sequence, was investigated using a fluorescence correlation spectroscopy (FCS). The transient dark state was measured in various solvents that are known to affect the intersystem crossing or photoisomerization of the DNA probe. On the basis of the experimental results, a simplified two energy state model of the probe was constructed, and this model provides an insight into the nonradiative relaxation mechanism of the fluorophore and the applications for DNA and RNA detection. The transient dark state that was measured in a time scale of a few microseconds is a triplet state and is related to photoisomerization, viscosity, oxygen concentration, and hybridization, all of which are important parameters for cellular microscopy. The transient dark state in a time scale of a sub-microseconds is sensitively changed after the addition of target DNA. The characterization can improve the probe’s capability to identify target DNA/RNA by using FCS since the triplet state that occurred after hybridization is distinctive in the time scale with that occurred before hybridization.