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微型化和自动化的自适应实验室进化:使谷氨酸棒杆菌朝着提高 d-木糖利用的方向进化。

Miniaturized and automated adaptive laboratory evolution: Evolving Corynebacterium glutamicum towards an improved d-xylose utilization.

机构信息

Institute of Bio- and Geosciences, IBG-1: Biotechnology, Forschungszentrum Jülich GmbH, Jülich D-52425, Germany.

Institute of Bio- and Geosciences, IBG-1: Biotechnology, Forschungszentrum Jülich GmbH, Jülich D-52425, Germany.

出版信息

Bioresour Technol. 2017 Dec;245(Pt B):1377-1385. doi: 10.1016/j.biortech.2017.05.055. Epub 2017 May 12.

Abstract

Adaptive Laboratory Evolution (ALE) is increasingly being used as a technique for untargeted strain optimization. This work aimed at developing an automated and miniaturized ALE approach based on repetitive batch cultivations in microtiter plates. The new method is applied to the recently published strain Corynebacterium glutamicum pEKEx3-xylXABCD, which is capable of utilizing d-xylose via the Weimberg (WMB) pathway. As a result, the significantly improved strain WMB2 was obtained, showing a specific growth rate of 0.26h on d-xylose as sole carbon and energy source. WMB2 grows stable during lab-scale bioreactor operation, demonstrating the high potential of this strain for future biorefinery applications. Genome sequencing of cell samples from two different ALE processes revealed potential key mutations, e.g. in the gene cg0196 (encoding for the transcriptional regulator IolR of the myo-inositol metabolism). These findings open up new perspectives for the rational engineering of C. glutamicum towards improved d-xylose utilization.

摘要

适应性实验室进化(ALE)越来越多地被用作非靶向菌株优化的技术。本工作旨在开发一种基于微滴定板重复分批培养的自动化和微型 ALE 方法。该新方法应用于最近发表的能够通过 Weimberg(WMB)途径利用 d-木糖的谷氨酸棒杆菌 pEKEx3-xylXABCD 菌株。结果,得到了显著改善的菌株 WMB2,在 d-木糖作为唯一碳源和能源时的比生长速率为 0.26h。WMB2 在实验室规模的生物反应器操作中稳定生长,表明该菌株在未来生物炼制应用中有很高的潜力。对来自两个不同 ALE 过程的细胞样品进行基因组测序揭示了潜在的关键突变,例如 cg0196 基因(编码肌醇代谢的转录调节剂 IolR)。这些发现为理性工程谷氨酸棒杆菌以提高 d-木糖利用开辟了新的视角。

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