Reduction of Ethyl Carbamate in an Alcoholic Beverage by CRISPR/Cas9-Based Genome Editing of the Wild Yeast

被引:10
作者
Jung, Jin-Young [1 ]
Kang, Min-Ji [1 ]
Hwang, Hye-Seon [1 ]
Baek, Kwang-Rim [1 ]
Seo, Seung-Oh [1 ]
机构
[1] Catholic Univ Korea, Dept Food Sci & Nutr, Bucheon 14662, South Korea
基金
新加坡国家研究基金会;
关键词
ethyl carbamate; alcoholic beverages; Saccharomyces cerevisiae; CRISPR; Cas9; genome editing; SACCHAROMYCES-CEREVISIAE; FERMENTATION; STRAINS; DELETION; MINIMIZE; CAR1;
D O I
10.3390/foods12010102
中图分类号
TS2 [食品工业];
学科分类号
0832 ;
摘要
Ethyl carbamate (EC) is a naturally occurring substance in alcoholic beverages from the reaction of ethanol with urea during fermentation and storage. EC can cause dizziness and vomiting when consumed in small quantities and develop kidney cancer when consumed in excess. Thus, the reduction of EC formation in alcoholic beverages is important for food safety and human health. One of the strategies for reducing EC contents in alcoholic beverages is developing a new yeast starter strain to enable less formation of EC during fermentation. In this study, we isolated a polyploid wild-type yeast Saccharomyces cerevisiae strain from the Nuruk (Korean traditional grain-based inoculum of wild yeast and mold) and developed a starter culture by genome engineering to reduce EC contents in alcoholic beverages. We deleted multiple copies of the target genes involved in the EC formation in the S. cerevisiae by a CRISPR/Cas9-based genome editing tool. First, the CAR1 gene encoding for the arginase enzyme responsible for the formation of urea was completely deleted in the genome of S. cerevisiae. Additionally, the GZF3 gene encoding the transcription factor controlling expression levels of several genes (DUR1, 2, and DUR3) related to urea absorption and degradation was deleted in S. cerevisiae to further reduce the EC formation. The effects of gene deletion were validated by RT-qPCR to confirm changes in transcriptional levels of the EC-related genes. The resulting strain of S. cerevisiae carrying a double deletion of CAR1 and GZF3 genes successfully reduced the EC contents in the fermentation medium without significant changes in alcohol contents and fermentation profiles when compared to the wild-type strain. Finally, we brewed the Korean traditional rice wine Makgeolli using the double deletion strain of S. cerevisiae dCAR1&GZF3, resulting in a significant reduction of the EC content in Makgeolli up to 41.6% when compared to the wild-type strain. This study successfully demonstrated the development of a starter culture to reduce the EC formation in an alcoholic beverage by CRISPR/Cas9 genome editing of the wild yeast.
引用
收藏
页数:13
相关论文
共 50 条
  • [41] An Efficient Genetic Transformation and CRISPR/Cas9-Based Genome Editing System for Moso Bamboo (Phyllostachys edulis)
    Huang, Biyun
    Zhuo, Renying
    Fan, Huijin
    Wang, Yujun
    Xu, Jing
    Jin, Kangming
    Qiao, Guirong
    FRONTIERS IN PLANT SCIENCE, 2022, 13
  • [42] CRISPR/Cas9-Based Genome Editing as a Way Ahead for Inducing Production of Bioactive Metabolites in Endophytes COMMENT
    Chowdhary, Kanika
    Arora, Himanshu
    Sharma, Satyawati
    NATIONAL ACADEMY SCIENCE LETTERS-INDIA, 2022, 45 (03): : 275 - 280
  • [43] First Report on CRISPR/Cas9-Based Genome Editing in the Destructive Invasive Pest Tuta Absoluta (Meyrick) (Lepidoptera: Gelechiidae)
    Ji, Shun-Xia
    Bi, Si-Yan
    Wang, Xiao-Di
    Wu, Qiang
    Tang, Yan-Hong
    Zhang, Gui-Fen
    Wan, Fang-Hao
    Lue, Zhi-Chuang
    Liu, Wan-Xue
    FRONTIERS IN GENETICS, 2022, 13
  • [44] Plasmid-free CRISPR/Cas9 genome editing in Saccharomyces cerevisiae
    Nishimura, Akira
    Tanahashi, Ryoya
    Oi, Tomoki
    Kan, Kyoyuki
    Takagi, Hiroshi
    BIOSCIENCE BIOTECHNOLOGY AND BIOCHEMISTRY, 2023, 87 (04) : 458 - 462
  • [45] CRISPR-Cas9 Based Bacteriophage Genome Editing
    Zhang, Xueli
    Zhang, Chaohui
    Liang, Caijiao
    Li, Bizhou
    Meng, Fanmei
    Ai, Yuncan
    MICROBIOLOGY SPECTRUM, 2022, 10 (04):
  • [46] Developing a CRISPR/Cas9 System for Genome Editing in the Basidiomycetous Yeast Rhodosporidium toruloides
    Jiao, Xiang
    Zhang, Yue
    Liu, Xiangjian
    Zhang, Qi
    Zhang, Sufang
    Zhao, Zongbao K.
    BIOTECHNOLOGY JOURNAL, 2019, 14 (07)
  • [47] Single-step Precision Genome Editing in Yeast Using CRISPR-Cas9
    Akhmetov, Azat
    Laurent, Jon M.
    Gollihar, Jimmy
    Gardner, Elizabeth C.
    Garge, Riddhiman K.
    Ellington, Andrew D.
    Kachroo, Aashiq H.
    Marcotte, Edward M.
    BIO-PROTOCOL, 2018, 8 (06):
  • [48] Increasing the efficiency of CRISPR/Cas9-based gene editing by suppressing RNAi in plants
    Xiaoyan Wang
    Jiayun Lu
    Kangwen Lao
    Suikang Wang
    Xiaowei Mo
    Xintong Xu
    Xuemei Chen
    Beixin Mo
    Science China(Life Sciences), 2019, (07) : 982 - 984
  • [49] A CRISPR/Cas9-based single-stranded DNA recombineering system for genome editing of Rhodococcus opacus PD630
    Liang, Youxiang
    Wei, Yuwen
    Jiao, Song
    Yu, Huimin
    SYNTHETIC AND SYSTEMS BIOTECHNOLOGY, 2021, 6 (03) : 200 - 208
  • [50] CRISPR-Cas9 Mediated Genome Editing in Drosophila
    Peng, Ping
    Wang, Xia
    Shen, Da
    Sun, Jin
    Jia, Yu
    Xu, Rong-Gang
    Zhu, Li-Fei
    Ni, Jian-Quan
    BIO-PROTOCOL, 2019, 9 (02):