RNA isolation from bacteria is technically difficult due to the RNA characteristic of labile and vulnerable degradation. Many reagents were explored for cellular lysis and complete inhibition of RNase. However, the available methods for RNA isolation are either of low efficiency or time-consuming. Here, we developed a rapid and accessible protocol for RNA isolation that combined a simplified cell lysis and RNA release by formamide-based solution and RNA purification by chitosan-modified silica membrane for the first time. With this method, we obtained about ~ 28 μg of total RNA from 10⁸ Escherichia coli cells. The entire procedure can be done within 15 min without redundant pipetting steps. The purity of extracted RNA was comparable to that of commercial kits, but the cost was much lower. Furthermore, the yielded RNA was successfully used in downstream enzymatic reactions, such as reverse transcription and quantitative real-time PCR. This new method would be of benefit for an extensive range of gene expression analyses in bacterial organisms.

Graphical abstract

中文翻译：

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通过整合甲酰胺提取和壳聚糖修饰的二氧化硅纯化从细菌中分离 RNA 的简单方法

由于 RNA 不稳定和易降解的特征，从细菌中分离 RNA 在技术上是困难的。探索了许多试剂用于细胞裂解和完全抑制 RNase。然而，现有的 RNA 分离方法要么效率低，要么耗时。在这里，我们开发了一种快速且易于使用的 RNA 分离方案，该方案首次结合了基于甲酰胺的溶液的简化细胞裂解和 RNA 释放以及壳聚糖改性硅胶膜的 RNA 纯化。使用这种方法，我们从 10 ⁸ 大肠杆菌中获得了约 28 μg 的总 RNA细胞。整个过程可在 15 分钟内完成，无需多余的移液步骤。提取的RNA纯度与商业试剂盒相当，但成本要低得多。此外，所产生的 RNA 已成功用于下游酶促反应，如逆转录和定量实时 PCR。这种新方法将有利于细菌生物体中广泛的基因表达分析。

图形概要