文献类型：期刊文献

中文题名：米曲霉产米黑根毛霉脂肪酶高通量筛选方法的建立

英文题名：Establishment of a High-Throughput Screening Method for Rhizomucor miehei Lipase Production by A.oryzae

作者：熊志岳[1];许逗[1];郭元昕[1];田锡炜[1];储炬[1]

机构：[1]华东理工大学生物反应器工程国家重点实验室,上海200237

年份：2022

卷号：48

期号：4

起止页码：519

中文期刊名：华东理工大学学报（自然科学版）

外文期刊名：Journal of East China University of Science and Technology

收录：Scopus;北大核心:【北大核心2020】；CSCD:【CSCD_E2021_2022】；

基金：国家重点研发计划(2019YFA0904803)。

语种：中文

中文关键词：米黑根毛霉脂肪酶;米曲霉;高通量筛选;ARTP

外文关键词：Rhizomucor miehei lipase(RML);Aspergillus oryzae;high-throughput screening;atmospheric room temperature plasma(ARTP)

摘要：为了快速、高效地筛选出高产米黑根毛霉脂肪酶(RML)的米曲霉菌株,利用常温常压等离子体诱变技术(ARTP)对米曲霉孢子进行诱变,优化了高通量检测RML酶活的双指示剂法,建立了24孔板米曲霉高通量培养体系。通过对诱变后的1200孔(36000~48000个孢子)进行高通量筛选,最终筛选出4株米曲霉高产突变株。经由摇瓶验证,突变株Ⅰ7-D6-7、Ⅱ4-4B-5、Ⅲ3-5C-1、Ⅴ7-6C-3酶活分别达到了265.98、240.90、253.52、293.50 U/mL,比出发菌株的酶活(176.52 U/mL)分别提高了50.68%、36.47%、43.61%、66.27%。
In order to quickly and efficiently screen out Aspergillus oryzae strains of high-yielding Rhizomucor miehei lipase(RML),the spores of Aspergillus oryzae(A.oryzae)were mutated by atmospheric room temperature plasma(ARTP)mutagenesis technology.The dual indicator method for high-throughput detection of RML enzyme activity was optimized,and a 24-well plate high-throughput culture system of A.oryzae was established.Through highthroughput screening of 1,200 wells(36,000-48,000 spores)after mutagenesis,4 high-yielding mutants,which was designatedⅠ7-D6-7,Ⅱ4-4B-5,Ⅲ3-5C-1 andⅤ7-6C-3,were finally obtained.In shake flask cultures,the enzyme activities of mutants reached 265.98,240.90,253.52 U/mL and 293.50 U/mL,which was increased by 50.68%,36.47%,43.61%and 66.27%,respectively,compared to the original strain(176.52 U/mL).