Muséum National d'Histoire Naturelle, CNRS, UMR 7196, INSERM, U565, 57 rue Cuvier, B.P. 26, Paris Cedex 05, F-75231, France.
Molecules. 2013 Dec 11;18(12):15357-97. doi: 10.3390/molecules181215357.
This review analyses the literature concerning non-fluorescent and fluorescent probes for nucleic acid imaging in fixed and living cells from the point of view of their suitability for imaging intracellular native RNA and DNA. Attention is mainly paid to fluorescent probes for fluorescence microscopy imaging. Requirements for the target-binding part and the fluorophore making up the probe are formulated. In the case of native double-stranded DNA, structure-specific and sequence-specific probes are discussed. Among the latest, three classes of dsDNA-targeting molecules are described: (i) sequence-specific peptides and proteins; (ii) triplex-forming oligonucleotides and (iii) polyamide oligo(N-methylpyrrole/N-methylimidazole) minor groove binders. Polyamides seem to be the most promising targeting agents for fluorescent probe design, however, some technical problems remain to be solved, such as the relatively low sequence specificity and the high background fluorescence inside the cells. Several examples of fluorescent probe applications for DNA imaging in fixed and living cells are cited. In the case of intracellular RNA, only modified oligonucleotides can provide such sequence-specific imaging. Several approaches for designing fluorescent probes are considered: linear fluorescent probes based on modified oligonucleotide analogs, molecular beacons, binary fluorescent probes and template-directed reactions with fluorescence probe formation, FRET donor-acceptor pairs, pyrene excimers, aptamers and others. The suitability of all these methods for living cell applications is discussed.
这篇综述从适用于细胞内天然 RNA 和 DNA 成像的角度分析了固定和活细胞中核酸成像的非荧光和荧光探针的文献。主要关注荧光显微镜成像的荧光探针。制定了构成探针的靶结合部分和荧光团的要求。对于天然双链 DNA,讨论了结构特异性和序列特异性探针。在最新的研究中,描述了三类靶向 dsDNA 的分子:(i)序列特异性肽和蛋白质;(ii)三链形成寡核苷酸;(iii)聚酰胺寡聚(N-甲基吡咯/N-甲基咪唑)小沟结合物。聚酰胺似乎是设计荧光探针最有前途的靶向试剂,但仍存在一些技术问题需要解决,例如相对较低的序列特异性和细胞内较高的背景荧光。引用了几个用于固定和活细胞中 DNA 成像的荧光探针应用的例子。对于细胞内 RNA,只有修饰的寡核苷酸才能提供这种序列特异性成像。考虑了几种设计荧光探针的方法:基于修饰的寡核苷酸类似物的线性荧光探针、分子信标、双荧光探针和荧光探针形成的模板指导反应、FRET 供体-受体对、芘激基缔合物、适体等。讨论了所有这些方法用于活细胞应用的适用性。