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Gross transcriptomic analysis of Pseudomonas putida for diagnosing environmental shifts.
Microbial Biotechnology ( IF 4.8 ) Pub Date : 2019-04-07 , DOI: 10.1111/1751-7915.13404
Ángeles Hueso-Gil 1 , Belén Calles 1 , George A O'Toole 2 , Víctor de Lorenzo 1
Affiliation  

The biological regime of Pseudomonas putida (and any other bacterium) under given environmental conditions results from the hierarchical expression of sets of genes that become turned on and off in response to one or more physicochemical signals. In some cases, such signals are clearly defined, but in many others, cells are exposed to a whole variety of ill‐defined inputs that occur simultaneously. Transcriptomic analyses of bacteria passed from a reference condition to a complex niche can thus expose both the type of signals that they experience during the transition and the functions involved in adaptation to the new scenario. In this article, we describe a complete protocol for generation of transcriptomes aimed at monitoring the physiological shift of P. putida between two divergent settings using as a simple case study the change between homogeneous, planktonic lifestyle in a liquid medium and growth on the surface of an agar plate. To this end, RNA was collected from P. putidaKT2440 cells at various times after growth in either condition, and the genome‐wide transcriptional outputs were analysed. While the role of individual genes needs to be verified on a case‐by‐case basis, a gross inspection of the resulting profiles suggested cells that are cultured on solid media consistently had a higher translational and metabolic activity, stopped production of flagella and were conspicuously exposed to a strong oxidative stress. The herein described methodology is generally applicable to other circumstances for diagnosing lifestyle determinants of interest.

中文翻译:

恶臭假单胞菌的总转录组学分析,用于诊断环境变化。

在给定的环境条件下,恶臭假单胞菌(和其他细菌)的生物学机制是由一组基因的分层表达引起的,这些基因根据一种或多种理化信号而开启和关闭。在某些情况下,此类信号已明确定义,但在其他情况下,细胞则暴露于多种同时发生的不确定输入中。从参考条件传递到复杂环境的细菌的转录组学分析可以揭示细菌在过渡过程中遇到的信号类型以及适应新情况的功能。在本文中,我们描述了一个完整的转录组生成协议,旨在监测恶臭假单胞菌的生理变化在两个不同的环境之间进行比较,以一个简单的案例为例,研究液体介质中均匀的浮游生活方式与琼脂平板表面的生长之间的变化。为此,在任一条件下生长后的不同时间,从恶臭假单胞菌KT2440细胞中收集RNA ,并对全基因组转录输出进行分析。虽然个别基因的作用需要根据具体情况进行验证,但对生成的图谱的粗略检查表明,在固体培养基上培养的细胞始终具有较高的翻译和代谢活性,停止了鞭毛的产生,并且明显地暴露于强烈的氧化应激中。本文描述的方法通常适用于诊断感兴趣的生活方式决定因素的其他情况。
更新日期:2019-04-07
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