Laboratory of Molecular Biology of Viruses, Belozersky Research Institute of Physico-Chemical Biology, Lomonosov Moscow State University, Moscow, Russian Federation; Laboratory of Immunochemistry, Research Center of Biotechnology, Russian Academy of Sciences, Moscow, Russian Federation.
Laboratory of Immunochemistry, Research Center of Biotechnology, Russian Academy of Sciences, Moscow, Russian Federation; Chair of Virology, Faculty of Biology, Lomonosov Moscow State University, Moscow, Russian Federation.
Anal Biochem. 2021 May 1;620:114113. doi: 10.1016/j.ab.2021.114113. Epub 2021 Jan 30.
The diversity of organisms, tissues and cells is so great that, to date, no universal method for RNA extraction from these biological materials exist. The RNA isolation technique with a mix of guanidine thiocyanate, phenol, and chloroform is most widely used. Extraction and purification of RNA methods using selling guanidinium-phenol (TRIzol)-based and silica-based column kits have limitations on toxicity, or RNA isolation, particularly for plants, and scaling. The agents' toxicity is particularly relevant when employing for mass analysis in practice while gaining RNA preparations during the pandemics, epizootics, and epiphytotic. In modern diagnostics of infections at the molecular level, powerful RT-PCR technology is used, which amplifies the detection of RNA pathogens by hundreds of millions of times. We proposed obtaining RNA samples from viruses, bacteria, and plants for the reverse transcription reactions with a subsequent amplification of cDNAs by the polymerase chain reaction using potent and nontoxic chaotropic agent ammonium trichloroacetate. The method works in the analytical and preparative range and can be useful in the case of extraordinary circumstances during mass infections. Potentially this method can be adapted for obtaining RNA samples ready for the RT-isothermal PCR in the field.
生物组织和细胞的多样性非常大，以至于迄今为止，还没有一种通用的方法可以从这些生物材料中提取 RNA。最广泛使用的是混合了硫氰酸胍、苯酚和氯仿的 RNA 提取技术。使用商品化的胍-酚（TRIzol）和基于硅胶的柱试剂盒提取和纯化 RNA 的方法，在毒性、RNA 分离方面存在限制，特别是对于植物和规模化方面。在大规模分析实践中，这些试剂的毒性尤其相关，而在大流行、动物流行病和植物流行病期间获得 RNA 制剂时更是如此。在分子水平上进行感染的现代诊断中，使用了强大的 RT-PCR 技术，该技术将 RNA 病原体的检测扩增了数亿倍。我们建议使用强毒性的三氯乙酸铵从病毒、细菌和植物中获得 RNA 样本，用于逆转录反应，随后通过聚合酶链反应扩增 cDNA。该方法适用于分析和制备范围，在大规模感染的特殊情况下可能非常有用。该方法可能适用于在野外获得准备用于 RT-等温 PCR 的 RNA 样本。
