Mondragon Paula, Hwang Sungmin, Schmid Amy, Maupin-Furlow Julie A
Department of Microbiology and Cell Science, Institute of Food and Agricultural Sciences, University of Florida, Gainesville, Florida, USA.
Department of Biology, Duke University, Durham, NC, 27708, USA.
Bio Protoc. 2022 Nov 20;12(22). doi: 10.21769/BioProtoc.4557.
The study of haloarchaea provides an opportunity to expand understanding of the mechanisms used by extremophiles to thrive in and respond to harsh environments, including hypersaline and oxidative stress conditions. A common strategy used to investigate molecular mechanisms of stress response involves the deletion and/or site-directed mutagenesis of genes identified through omics studies followed by a comparison of the mutant and wild-type strains for phenotypic differences. The experimental methods used to monitor these differences must be controlled and reproducible. Current methods to examine recovery of halophilic archaea from extreme stress are complicated by extended incubation times, nutrients not typically encountered in the environment, and other related limitations. Here we describe a method for assessing the function of genes during hypochlorite stress in the halophilic archaeon that overcomes these types of limitations. The method was found reproducible and informative in identifying genes needed for to recover from hypochlorite stress.
嗜盐古菌的研究为扩展对极端微生物在包括高盐和氧化应激条件等恶劣环境中生存及响应所采用机制的理解提供了契机。用于研究应激反应分子机制的一种常见策略是对通过组学研究鉴定出的基因进行缺失和/或定点诱变，随后比较突变株和野生型菌株的表型差异。用于监测这些差异的实验方法必须可控且可重复。目前用于检测嗜盐古菌从极端应激中恢复的方法因孵育时间延长、环境中不常见的营养物质以及其他相关限制而变得复杂。在此，我们描述了一种在嗜盐古菌次氯酸盐应激期间评估基因功能的方法，该方法克服了这类限制。该方法在鉴定从次氯酸盐应激中恢复所需的基因方面具有可重复性且信息丰富。
