Department of Medical Laboratory Technology, Bapubhai Desaibhai Patel Institue of Paramedical Sciences (BDIPS), Charotar University of Science and Technology, CHARUSAT campus, Changa, Anand, Gujarat, 388421, India.
Bapubhai Desaibhai Patel Institute of Paramedical Sciences, Charotar University of Science and Technology, Anand, India.
Arch Microbiol. 2024 Aug 7;206(9):369. doi: 10.1007/s00203-024-04077-2.
The RNA-based study provides an excellent indication of an organism's gene expression profile. Obtaining high-yield and high-purity RNA from Gram-positive and acid-fast bacteria is difficult without high-end kits and facilities. We optimised effective and simple protocol for RNA isolation that is a combination of enzymatic, physical and chemical treatment to disrupt cells. We successfully isolated high quality intact total RNA with yields ranging from 23.13 ± 0.40 to 61.51 ± 0.27 µg and the 260/280 purity ratio of 1.95 ± 0.01 to 2.05 ± 0.01 from Staphylococcus aureus, Staphylococcus epidermidis, Enterococcus faecalis, and Mycobacterium smegmatis. These results represents a significantly enhanced yield and purity compared to other combination of techniques which we performed. Compared to previous studies the yield obtained by this method is high for the studied organisms. Furthermore the yielded RNA was successfully used for downstream applications such as quantitative real time PCR. The described method can be easily optimised and used for various bacteria.
基于 RNA 的研究为生物体的基因表达谱提供了很好的指示。如果没有高端试剂盒和设备，从革兰氏阳性菌和抗酸性菌中获得高产率和高纯度的 RNA 是很困难的。我们优化了一种有效的简单 RNA 分离方案，该方案结合了酶、物理和化学处理，以破坏细胞。我们成功地从金黄色葡萄球菌、表皮葡萄球菌、粪肠球菌和耻垢分枝杆菌中分离出了高质量的完整总 RNA，产量范围为 23.13±0.40 至 61.51±0.27µg，260/280 纯度比为 1.95±0.01 至 2.05±0.01。与我们之前进行的其他技术组合相比，这些结果显示出明显更高的产量和纯度。与之前的研究相比，该方法获得的研究生物的产量很高。此外，所得到的 RNA 可成功用于下游应用，如定量实时 PCR。所描述的方法可以很容易地进行优化，并用于各种细菌。
