嗜热毁丝霉高通量培养技术及其应用

A high-throughput cultivation technology and its application for Myceliophthora thermophila

摘要:

【背景】丝状真菌是一类重要的工业发酵生产宿主菌，如何进行高通量纯菌培养和高效检测筛选性能优异的菌株是工业丝状真菌研究的重要方向。【目的】研究建立丝状真菌的高通量培养技术并测试应用效果。【方法】通过对丝状真菌培养过程中的制种、接种、培养和检测研究，建立基于孔板的高通量培养技术，并以嗜热毁丝霉为例对该技术进行验证。【结果】与传统的平板制种和摇瓶接种培养方式相比，高通量孔板的培养方式将制种通量提高24倍，单位面积产孢子能力提高350%，液体培养转接效率提高10−40倍，并建立96孔板测定乙醇含量的高通量检测技术。【结论】将丝状真菌的培养和检测通量提高1−2个数量级，为快速检测丝状真菌改造过程产生的大量性状不同菌株并获得目标菌株奠定基础，为丝状真菌高通量筛选研究提供应用指导价值。

Abstract:

[Background] Filamentous fungi are a key group of host fungi in industrial fermentation. How to carry out high-throughput cultivation and screen out the efficient strains is an important direction of the research on industrial filamentous fungi. [Objective] To establish a high-throughput cultivation technology of filamentous fungi and evaluate its efficiency. [Methods] After studying the seed production, inoculation, cultivation, and detection of filamentous fungi, we established a microplate-based high-throughput cultivation technology and evaluated its performance with Myceliophthora thermophila. [Results] Compared with the traditional methods using either plate or shake flask for seed production, the microplate-based method increased the seed production by 24 folds, the spore production per unit area by 350%, and the transfer efficiency of liquid culture by 10-40 folds. Furthermore, a 96-well microplate-based high-throughput technique was developed for ethanol content determination. [Conclusion] The developed high-throughput technology increased the cultivation and detection efficiency of filamentous fungi by 1−2 orders of magnitude. This study lays a foundation for rapid screening of the target strains from a large number of variants with different traits and provides a reference for the research on filamentous fungi.