ARTP-MMC高通量筛选耐木质纤维素预处理抑制物酿酒酵母的研究  

High-throughput screening of inhibitors tolerance Saccharomyces cerevisiae from lignocellulosic pretreatment by ARTP-MMC

作　　者：古永红[1] 付少虎 廖蓓[3] 龚大春 吕育财 王昕 李志军[3] GU Yonghong;FU Shaohu;LIAO Bei;GONG Dachun;LV Yucai;WANG Xin;LI Zhijun(College of Mechanical and Electrical Engineering,Hubei Three Gorges Polytechnic,Yichang 443000,China;Colege of Biological and Pharmaceutical,China Three Gorges University,Yichang 443002,China;Angel Yeast Co,Ld,Yichang443003,China)

机构地区：[1]湖北三峡职业技术学院机电工程学院,湖北宜昌443000 [2]三峡大学生物与制药学院,湖北宜昌443002 [3]安琪酵母股份有限公司,湖北宜昌443003

出　　处：《中国酿造》2024年第9期99-104,共6页China Brewing

基　　金：国家自然基金面上项目(21776162)。

摘　　要：为了提升酿酒酵母(Saccharomyces cerevisiae)对木质纤维素预处理所产生的甲酸、糠醛、5-羟甲基糠醛等抑制物的耐受性,保障酿酒酵母产乙醇能力不受影响,该文以安琪超级酿酒酵母为出发菌株,研究了一种常温常压等离子体(ARTP)诱变和微液滴细胞培养(MMC)相结合的适应性进化和高通量筛选耐木质纤维素预处理抑制物的酿酒酵母方法,获得抗性强的酿酒酵母优势突变株,并评价其耐抑制物、耐高温和产乙醇能力。结果表明,菌株最佳诱变条件为:诱变功率110 W,诱变时间140 s,培养12 h,微液滴微生物培养的适应性进化复合抑制剂梯度组合为CI4~CI6,经过高通量筛选获得最佳的突变株M8;该菌株可耐受体积分数16%乙醇,具有较好的耐高温特性,55℃热激5 min可正常生长;经过10 L发酵罐培养,其最大乙醇产量为56.29 g/L,糖醇转化率为0.49 g/g,达到理论值的96.67%。该研究对木质纤维素高质化利用提供了优良的酿酒酵母,对推动纤维素乙醇具有重要意义。In order to enhance the tolerance of Saccharomyces cerevisiae to inhibitors such as formic acid,furfural,5-hydroxymethyl-furfural produced by lignocellulosic pretreatment,and to ensure that the ethanol production capacity of S.cerevisiae was not affected,using Angel's super yeast as the starting strain,an adaptive evolution method combining atmospheric roomtemperature plasma(ARTP)mutagenesis and microbial microdroplets culture(MMC)with high throughput screening of lignocellulose-resistant preconditioning inhibitors for S.cerevisiae was studied.a dominate mutant strain with strong resistance was obtained and its inhibition resistance and ethanol production capacity were evaluated.The research showed that the optimal mutation conditions for the strain were as follows:mutation power 110 W,mutation time 140 s,cultivation time 12 h,and the compound inhibitors gradient combination of CI4-CI6 for adaptive evolution in the micro-droplet microbial culture system.So the optimal mutant strain M8 was obtained through high-throughput screening.The strain could tolerate 16%ethanol,had good high temperature resistance,and could grow normally at 55℃heat shock for 5 min.After 10 L fermenter culture,the maximum ethanol yield was 56.29 g/L,and the sugar-alcohol conversion rate was 0.49 g/g,reaching 96.67%of the theoretical value.The study provided excellent S.cerevisiae for the high-quality utilization of lignocellulosic biomass and was of great significance for promoting cellulose ethanol production.

关 键 词：常温常压等离子体诱变 微液滴细胞培养 高通量筛选 耐抑制物 酿酒酵母 

分 类 号：TK6[动力工程及工程热物理—生物能] Q813[生物学—生物工程]