Single-cell detection of microRNAs in developing vertebrate embryos after acute administration of a dual-fluorescence reporter/sensor plasmid
Single-cell detection of microRNAs in developing vertebrate embryos after acute administration of a dual-fluorescence reporter/sensor plasmid
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DOI：
10.2144/000112296
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发表时间：
2006-12-01
期刊：
影响因子：
2.7
通讯作者：
Huttner, Wieland B.
中科院分区：
文献类型：
--
作者：
Tonelli, Davide De Pietri;Calegari, Federico;Huttner, Wieland B.
单细胞分辨率的 microRNA (miRNA) 检测对于研究这些转录后调节因子的作用非常重要。在这里，我们使用双荧光绿色荧光蛋白（GFP）-报告基因/单体红色荧光蛋白（mRFP）-传感器（DFRS）质粒，将其注射到斑马鱼卵裂球中或电穿孔到子宫内或子宫外小鼠胚胎的特定组织中，以监测单个活细胞中特定miRNA的动态。例如，这种方法揭示，在发育中的小鼠中枢神经系统中，miR-124a 不仅在有丝分裂后神经元中表达，而且在神经元祖细胞中表达。总的来说，我们的结果表明，DFRS 质粒的急性施用提供了先前原位杂交和转基因方法的替代方案，并允许在脊椎动物发育过程中监测特定细胞谱系中 miRNA 的出现和消失。
The detection of microRNAs (miRNAs) at single-cell resolution is important for studying the role of these posttranscriptional regulators. Here, we use a dual-fluorescent green fluorescent protein (GFP)-reporter/monomeric red fluorescent protein (mRFP)-sensor (DFRS) plasmid, injected into zebrafish blastomeres or electroporated into defined tissues of mouse embryos in utero or ex utero, to monitor the dynamics of specific miRNAs in individual live cells. This approach reveals, for example, that in the developing mouse central nervous system,, miR-124a is expressed not only in postmitotic neurons but also in neuronal progenitor cells. Collectively, our results demonstrate that acute administration of DFRS plasmids.offers an alternative to previous in situ hybridization and transgenic approaches and allows the monitoring of miRNA appearance and disappearance in defined cell lineages during vertebrate development.