Institute for Integrated Cell-Material Sciences, Kyoto University, Kyoto 606-8501, Japan.
RNA. 2012 Jan;18(1):166-75. doi: 10.1261/rna.028431.111. Epub 2011 Nov 18.
Fluorescence in situ hybridization (FISH) is a powerful tool used in karyotyping, cytogenotyping, cancer diagnosis, species specification, and gene-expression analysis. Although widely used, conventional FISH protocols are cumbersome and time consuming. We have now developed a FISH method using exciton-controlled hybridization-sensitive fluorescent oligodeoxynucleotide (ECHO) probes. ECHO-FISH uses a 25-min protocol from fixation to mounting that includes no stringency washing steps. We use ECHO-FISH to detect both specific DNA and RNA sequences with multicolor probes. ECHO-FISH is highly reproducible, stringent, and compatible with other fluorescent cellular labeling techniques. The resolution allows detection of intranuclear speckles of poly(A) RNA in HeLa cells and dissociated hippocampal primary cultures, and mRNAs in the distal dendrites of hippocampal neurons. We also demonstrate detection of telomeric and centromeric DNA on metaphase mouse chromosomes. The simplicity of the ECHO-FISH method will likely accelerate cytogenetic and gene-expression analysis with high resolution.
荧光原位杂交（FISH）是一种用于核型分析、细胞遗传学分析、癌症诊断、物种鉴定和基因表达分析的强大工具。尽管广泛应用，但传统的 FISH 方案繁琐且耗时。我们现在已经开发出一种使用激子控制杂交敏感荧光寡脱氧核苷酸（ECHO）探针的 FISH 方法。ECHO-FISH 使用从固定到安装的 25 分钟方案，其中不包括严格的洗涤步骤。我们使用 ECHO-FISH 用多色探针检测特定的 DNA 和 RNA 序列。ECHO-FISH 具有高度可重复性、严格性，并且与其他荧光细胞标记技术兼容。该分辨率允许检测 HeLa 细胞和分离的海马原代培养物中核内多聚（A）RNA 的斑点以及海马神经元远端树突中的 mRNA。我们还证明了在中期小鼠染色体上检测端粒和着丝粒 DNA。ECHO-FISH 方法的简单性可能会加速具有高分辨率的细胞遗传学和基因表达分析。
