Institute for Sustainable Sciences and Development, Hiroshima University , Higashihiroshima, Hiroshima 739-8511, Japan.
Department of Molecular Biotechnology, Graduate School of Advanced Sciences of Matter, Hiroshima University , Higashihiroshima, Hiroshima 739-8530, Japan.
Anal Chem. 2016 Aug 16;88(16):7894-8. doi: 10.1021/acs.analchem.6b02710. Epub 2016 Jul 28.
This letter discusses the feasibility of continuously monitoring specific mRNA expression responses in a living cell with a probe structured as a fluorescence resonance energy transfer (FRET)-based DNA nano-tweezer (DNA-NT). The FRET-based DNA-NT, self-assembled from three single-stranded DNAs, alters its structure from an open state to a closed state in recognition of a target mRNA, resulting in the closing of the distal relation of previously modified FRET-paired fluorescent dyes and generating a FRET signal. The expressions of glucose transporters (GLUT) 1 and 4 in a mouse hepato-carcinoma (Hepa 1-6 cells) were selected as the target model. Live-cell imaging analysis of Hepa 1-6 cells with both FRET-based DNA-NTs indicated that the behaviors of the FRET signals integrated in each individual cell were similar to those measured with the conventional mass analysis technique of semiquantitative real-time (RT) polymerase chain reaction (PCR). From these results, it is concluded that continuous monitoring of gene expression response without gene recombination is feasible with a FRET-based DNA-NT, even in a single cell manner.
这封信讨论了使用荧光共振能量转移（FRET）为基础的 DNA 纳米夹（DNA-NT）作为探针，连续监测活细胞中特定 mRNA 表达反应的可行性。该 FRET 为基础的 DNA-NT 由三个单链 DNA 自组装而成，在识别靶 mRNA 时，其结构从开环状态转变为闭环状态，导致先前修饰的 FRET 配对荧光染料的远端关系关闭，并产生 FRET 信号。葡萄糖转运蛋白（GLUT）1 和 4 在小鼠肝癌（Hepa 1-6 细胞）中的表达被选为靶模型。用 FRET 为基础的 DNA-NT 对 Hepa 1-6 细胞进行的活细胞成像分析表明，整合在每个单个细胞中的 FRET 信号的行为与用传统的半定量实时（RT）聚合酶链反应（PCR）的质量分析技术测量的行为相似。从这些结果可以得出结论，即使在单细胞方式下，使用 FRET 为基础的 DNA-NT 也可以实现无需基因重组的基因表达响应的连续监测。
