SIGHT—A System for Solvent‐Tight Incubation and Growth Monitoring in High Throughput

IF 3.9 4区 生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY
Jakob Rönitz, Felix Herrmann, Benedikt Wynands, Tino Polen, Nick Wierckx
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引用次数: 0

Abstract

Physiological characterization of microorganisms in the context of solvent tolerance is a tedious process with a high investment of manual labor while often being limited in throughput capability simultaneously. Therefore, we developed a small‐scale solvent‐impervious cultivation system consisting of screw cap‐sealed glass vials in combination with a 3D‐printed vial holder for the Growth Profiler (EnzyScreen) platform. Components and cultivation conditions were empirically tested, and a suitable setup was found for the intended application. To demonstrate the capability of this cultivation system, an adaptive laboratory evolution was performed to further increase the tolerance of Pseudomonas taiwanensis GRC3 toward styrene. This approach yielded heterogenic cultures with improved growth performances in the presence of styrene from which individual clones were isolated and characterized in high throughput. Several clones with improved growth in the presence of 1% (v/v) styrene were analyzed through whole‐genome sequencing, revealing mutations in the co‐chaperone‐encoding gene dnaJ, RNA polymerase α subunit‐encoding gene rpoA, and loss‐of‐function mutations in the ttgGHI solvent efflux pump repressor encoded by ttgV. The developed cultivation system has proven to be a very useful extension of the Growth Profiler, as it reduces manual workload and allows high‐throughput characterization.
SIGHT--用于高通量溶剂密闭培养和生长监控的系统
耐溶剂微生物的生理学特征描述是一个繁琐的过程,需要投入大量的人工,同时通量能力往往有限。因此,我们开发了一种小规模不透溶剂培养系统,该系统由螺旋盖密封玻璃瓶和三维打印瓶架组成,适用于生长曲线仪(EnzyScreen)平台。根据经验对组件和培养条件进行了测试,找到了适合预期应用的设置。为了证明该培养系统的能力,进行了适应性实验室进化,以进一步提高台湾假单胞菌 GRC3 对苯乙烯的耐受性。这种方法产生了在苯乙烯存在下具有更好生长性能的异源培养物,从中分离出了单个克隆,并对其进行了高通量表征。通过全基因组测序分析了几个在 1%(v/v)苯乙烯存在下生长性能有所改善的克隆,发现了共伴侣蛋白编码基因 dnaJ、RNA 聚合酶 α 亚基编码基因 rpoA 的突变,以及由 ttgV 编码的 ttgGHI 溶剂外排泵抑制因子的功能缺失突变。事实证明,所开发的培养系统是生长曲线仪的一个非常有用的扩展,因为它减少了人工工作量,并能进行高通量表征。
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来源期刊
Engineering in Life Sciences
Engineering in Life Sciences 工程技术-生物工程与应用微生物
CiteScore
6.40
自引率
3.70%
发文量
81
审稿时长
3 months
期刊介绍: Engineering in Life Sciences (ELS) focuses on engineering principles and innovations in life sciences and biotechnology. Life sciences and biotechnology covered in ELS encompass the use of biomolecules (e.g. proteins/enzymes), cells (microbial, plant and mammalian origins) and biomaterials for biosynthesis, biotransformation, cell-based treatment and bio-based solutions in industrial and pharmaceutical biotechnologies as well as in biomedicine. ELS especially aims to promote interdisciplinary collaborations among biologists, biotechnologists and engineers for quantitative understanding and holistic engineering (design-built-test) of biological parts and processes in the different application areas.
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