Institute of Molecular Genetics, Academy of Sciences of the Czech Republic, Videnska 1083, 142 20 Prague 4, Czech Republic.
Methods. 2011 Feb;53(2):136-41. doi: 10.1016/j.ymeth.2010.08.005. Epub 2010 Aug 12.
RNA molecules rarely function alone in cells. For most RNAs, their function requires formation of various ribonucleoprotein (RNP) complexes. For example, mRNP composition can determine mRNA localization, translational repression, level of translation or mRNA stability. RNPs are usually studied by biochemical methods. However, biochemical approaches are unsuitable for some model systems, such as mammalian oocytes and early embryos, due to the small amounts that can be obtained for experimental analysis. In such cases, microscopic techniques are often used to learn about RNPs. Here, we present a review of immunostaining, fluorescence in situ hybridization with subcellular resolution and a combination of both, with emphasis on the mouse oocyte and early embryos models. Application of these techniques to whole-mount fixed oocytes and early embryos can provide information about RNP composition and localization with three-dimensional resolution.
RNA 分子在细胞中很少单独发挥作用。对于大多数 RNA 而言,其功能需要形成各种核糖核蛋白 (RNP) 复合物。例如,mRNA 的组成可以决定 mRNA 的定位、翻译抑制、翻译水平或 mRNA 的稳定性。RNP 通常通过生化方法进行研究。然而,由于可用于实验分析的量很少,生化方法不适用于某些模型系统,例如哺乳动物卵母细胞和早期胚胎。在这种情况下,通常使用显微镜技术来了解 RNP。在这里,我们回顾了免疫染色、亚细胞分辨率的荧光原位杂交以及两者的结合,重点介绍了小鼠卵母细胞和早期胚胎模型。这些技术在整个固定卵母细胞和早期胚胎上的应用可以提供有关 RNP 组成和定位的三维分辨率信息。