常压室温等离子体诱变与微生物微液滴培养选育谷胱甘肽高产菌株  

Breeding of a High-Yield Glutathione-Producing Strain by Atmospheric and Room Temperature Plasma Mutagenesis and Microbial Microdroplet Culture System

作　　者：凌思雨 王洲[1] 张会敏 李闯 刘庆涛 刘艳[1] 薛正莲[1] LING Siyu;WANG Zhou;ZHANG Huimin;LI Chuang;LIU Qingtao;LIU Yan;XUE Zhenglian(Anhui Engineering Laboratory for Industrial Microbiology Molecular Breeding,Anhui Engineering Technology Research Center of Microbial Fermentation,College of Biology and Food Engineering,Anhui Polytechnic University,Wuhu 241000,China)

机构地区：[1]安徽工程大学生物与食品工程学院,安徽省工业微生物分子育种工程实验室,微生物发酵安徽省工程技术研究中心,安徽芜湖241000

出　　处：《食品科学》2023年第4期200-208,共9页Food Science

基　　金：安徽省高校自然科学研究项目(KJ2017A120)。

摘　　要：为提升野生毕赤酵母菌株BY-1产谷胱甘肽的能力,通过常压室温等离子体(atmospheric and room temperature plasma,ARTP)诱变技术得到诱变菌株BY-1-26。进一步在摇瓶培养过程中添加1,2,4-三氮唑,提高诱变菌株产量。最后将1,2,4-三氮唑作为筛选因子,利用微生物微液滴培养(microbial microdroplet culture system,MMC)仪对诱变菌株进行适应性进化,获得了1株高产谷胱甘肽的突变株BY-2-24,并对其遗传稳定性进行研究。结果表明:出发菌株BY-1经过ARTP诱变处理、抗性梯度平板初筛、MMC适应性进化、摇瓶复筛等,可以选育高产谷胱甘肽突变株。突变株BY-2-24摇瓶产量达(312.13±2.62)mg/L,较出发菌株提高134.26%,且经过7次传代培养,仍然具有较好的遗传稳定性。同时,生物量提高118.33%,表明诱变菌株的生长能力得到提高。研究表明,ARTP与MMC联合应用作为一种简便高效的微生物诱变方式,可用于定向诱变筛选高性能微生物菌株,为高通量选育目标菌株提供了参考。In order to improve the ability of a wild-type strain(BY-1) of Pichia pastoris to produce glutathione(GSH),a mutant strain designated BY-1-26 was obtained by atmospheric and room temperature plasma(ARTP) mutagenesis.To further improve the production of glutathione by the mutant during shake flask culture,1,2,4-triazole was added to the culture medium.Finally,adaptive evolution of the mutant strain was carried out on a microbial microdroplet culture system(MMC) using 1,2,4-triazole as a screening factor.As a result,a high-yield glutathione-producing mutant strain BY-2-24 was obtained,and its genetic stability was investigated.The results showed that a high-yield glutathione-producing mutant strain could be obtained using the wild-type strain BY-1 by ARTP mutagenesis,primary resistance screening on gradient plates,adaptive evolution on MMC and shake-flask secondary screening.The yield of glutathione produced by BY-2-24 in shake flask culture was(312.13 ± 2.62) mg/L,which was 134.26% higher than that produced by the original strain,and this mutant still had good genetic stability after seven passages.Meanwhile,the biomass was increased by 118.33%,indicating that the growth ability of the mutant strain was improved compared to the original strain.The result of this study show that the combination of ARTP and MMC can be used as a simple and effective screening method for excellent mutant strains,which provides a reference for high throughput selection of target strains.

关 键 词：谷胱甘肽 毕赤酵母 常压室温等离子体诱变 适应性进化 菌种筛选 

分 类 号：Q815[生物学—生物工程]