Laboratory of Molecular Analysis, Department of Life Sciences, Federal University of Tocantins, Palmas, University Campus of Palmas, TO, Brazil.
Laboratory of Biotechnology, Food analysis, and Product Purification, Federal University of Tocantins, University Campus of Gurupi, TO, Brazil.
J Microbiol Methods. 2021 May;184:106200. doi: 10.1016/j.mimet.2021.106200. Epub 2021 Mar 10.
Isolating high quality RNA is a limiting factor in molecular analysis, since it is the base for transcriptional studies. The RNA extraction method can directly affect the RNA quality and quantity, as well as, its overall cost. The industrial importance of the yeast genus Candida in several sectors comes from their capacity to produce Lipases. These enzymes are one of the main metabolites produced by some Candida species, and it has been shown that Candida yeast can biodegrade petroleum hydrocarbons and diesel oil from biosurfactants that they can produce, a feature that turns these organisms into potential combatants for bioremediation techniques. Thus, this study aimed to determine an efficient method for isolating high quality RNA from Candida viswanathii biomass. To achieve this aim, three different RNA extraction methods, TRIzol, Hot Acid Phenol, and CTAB (Cetyltrimethylammonium Bromide), were tested. The three tested methods allowed the isolation of high-quality RNA from C. viswanathii biomass and yielded suitable RNA quantity for carrying out RT-qPCR studies. In addition, all methods displayed high sensitivity for the expression analysis of the CvGPH1 gene through RT-qPCR, with TRIzol and CTAB showing the best results and the CTAB method displaying the best cost-benefit ratio (US$0.35/sample).
从生物质中分离高质量 RNA 是分子分析的一个限制因素，因为它是转录研究的基础。RNA 提取方法直接影响 RNA 的质量和数量，以及其总体成本。酵母属 Candida 在几个领域的工业重要性来自于它们产生脂肪酶的能力。这些酶是一些 Candida 物种产生的主要代谢物之一，已经表明 Candida 酵母可以从它们可以产生的生物表面活性剂中生物降解石油烃和柴油，这一特性使这些生物体成为生物修复技术的潜在竞争者。因此，本研究旨在确定一种从 Candida viswanathii 生物质中分离高质量 RNA 的有效方法。为了实现这一目标，测试了三种不同的 RNA 提取方法：TRIzol、热酸苯酚和 CTAB（十六烷基三甲基溴化铵）。这三种方法均能从 C. viswanathii 生物质中分离出高质量的 RNA，并且能够获得足够的 RNA 量来进行 RT-qPCR 研究。此外，所有方法在通过 RT-qPCR 对 CvGPH1 基因的表达分析中都显示出了很高的灵敏度，其中 TRIzol 和 CTAB 方法的效果最好，而 CTAB 方法的成本效益比（每样 0.35 美元）最高。
