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Improving freeze-tolerance of baker's yeast through seamless gene deletion of NTH1 and PUT1.
J Ind Microbiol Biotechnol 2016; 43(6):817-28JI

Abstract

Baker's yeast strains with freeze-tolerance are highly desirable to maintain high leavening ability after freezing. Enhanced intracellular concentration of trehalose and proline in yeast is linked with freeze-tolerance. In this study, we constructed baker's yeast with enhanced freeze-tolerance by simultaneous deletion of the neutral trehalase-encoded gene NTH1 and the proline oxidase-encoded gene PUT1. We first used the two-step integration-based seamless gene deletion method to separately delete NTH1 and PUT1 in haploid yeast. Subsequently, through two rounds of hybridization and sporulation-based allelic exchange and colony PCR-mediated tetrad analysis, we obtained strains with restored URA3 and deletion of NTH1 and/or PUT1. The resulting strain showed higher cell survival and dough-leavening ability after freezing compared to the wild-type strain due to enhanced accumulation of trehalose and/or proline. Moreover, mutant with simultaneous deletion of NTH1 and PUT1 exhibits the highest relative dough-leavening ability after freezing compared to mutants with single-gene deletion perhaps due to elevated levels of both trehalose and proline. These results verified that it is applicable to construct frozen dough baker's yeast using the method proposed in this paper.

Authors+Show Affiliations

Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, Tianjin Industrial Microbiology Key Laboratory, College of Biotechnology, Tianjin University of Science and Technology, No. 29, 13th Avenue, Tianjin Economic and Technological Development Area, Tianjin, 300457, China. Tianjin Food Safety and Low Carbon Manufacturing Collaborative Innovation Center, Tianjin, 300457, People's Republic of China.Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, Tianjin Industrial Microbiology Key Laboratory, College of Biotechnology, Tianjin University of Science and Technology, No. 29, 13th Avenue, Tianjin Economic and Technological Development Area, Tianjin, 300457, China. Tianjin Food Safety and Low Carbon Manufacturing Collaborative Innovation Center, Tianjin, 300457, People's Republic of China.Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, Tianjin Industrial Microbiology Key Laboratory, College of Biotechnology, Tianjin University of Science and Technology, No. 29, 13th Avenue, Tianjin Economic and Technological Development Area, Tianjin, 300457, China.Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, Tianjin Industrial Microbiology Key Laboratory, College of Biotechnology, Tianjin University of Science and Technology, No. 29, 13th Avenue, Tianjin Economic and Technological Development Area, Tianjin, 300457, China. Tianjin Food Safety and Low Carbon Manufacturing Collaborative Innovation Center, Tianjin, 300457, People's Republic of China.Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, Tianjin Industrial Microbiology Key Laboratory, College of Biotechnology, Tianjin University of Science and Technology, No. 29, 13th Avenue, Tianjin Economic and Technological Development Area, Tianjin, 300457, China. Tianjin Food Safety and Low Carbon Manufacturing Collaborative Innovation Center, Tianjin, 300457, People's Republic of China.Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, Tianjin Industrial Microbiology Key Laboratory, College of Biotechnology, Tianjin University of Science and Technology, No. 29, 13th Avenue, Tianjin Economic and Technological Development Area, Tianjin, 300457, China. Tianjin Food Safety and Low Carbon Manufacturing Collaborative Innovation Center, Tianjin, 300457, People's Republic of China.Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, Tianjin Industrial Microbiology Key Laboratory, College of Biotechnology, Tianjin University of Science and Technology, No. 29, 13th Avenue, Tianjin Economic and Technological Development Area, Tianjin, 300457, China. Tianjin Food Safety and Low Carbon Manufacturing Collaborative Innovation Center, Tianjin, 300457, People's Republic of China.Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, Tianjin Industrial Microbiology Key Laboratory, College of Biotechnology, Tianjin University of Science and Technology, No. 29, 13th Avenue, Tianjin Economic and Technological Development Area, Tianjin, 300457, China. xdg@tust.edu.cn. Tianjin Food Safety and Low Carbon Manufacturing Collaborative Innovation Center, Tianjin, 300457, People's Republic of China. xdg@tust.edu.cn.

Pub Type(s)

Journal Article

Language

eng

PubMed ID

26965428

Citation

Dong, Jian, et al. "Improving Freeze-tolerance of Baker's Yeast Through Seamless Gene Deletion of NTH1 and PUT1." Journal of Industrial Microbiology & Biotechnology, vol. 43, no. 6, 2016, pp. 817-28.
Dong J, Chen D, Wang G, et al. Improving freeze-tolerance of baker's yeast through seamless gene deletion of NTH1 and PUT1. J Ind Microbiol Biotechnol. 2016;43(6):817-28.
Dong, J., Chen, D., Wang, G., Zhang, C., Du, L., Liu, S., ... Xiao, D. (2016). Improving freeze-tolerance of baker's yeast through seamless gene deletion of NTH1 and PUT1. Journal of Industrial Microbiology & Biotechnology, 43(6), pp. 817-28. doi:10.1007/s10295-016-1753-7.
Dong J, et al. Improving Freeze-tolerance of Baker's Yeast Through Seamless Gene Deletion of NTH1 and PUT1. J Ind Microbiol Biotechnol. 2016;43(6):817-28. PubMed PMID: 26965428.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Improving freeze-tolerance of baker's yeast through seamless gene deletion of NTH1 and PUT1. AU - Dong,Jian, AU - Chen,Didi, AU - Wang,Guanglu, AU - Zhang,Cuiying, AU - Du,Liping, AU - Liu,Shanshan, AU - Zhao,Yu, AU - Xiao,Dongguang, Y1 - 2016/03/10/ PY - 2015/11/28/received PY - 2016/02/16/accepted PY - 2016/3/12/entrez PY - 2016/3/12/pubmed PY - 2017/11/29/medline KW - Baker’s yeast KW - Proline KW - Seamless gene deletion KW - Trehalose SP - 817 EP - 28 JF - Journal of industrial microbiology & biotechnology JO - J. Ind. Microbiol. Biotechnol. VL - 43 IS - 6 N2 - Baker's yeast strains with freeze-tolerance are highly desirable to maintain high leavening ability after freezing. Enhanced intracellular concentration of trehalose and proline in yeast is linked with freeze-tolerance. In this study, we constructed baker's yeast with enhanced freeze-tolerance by simultaneous deletion of the neutral trehalase-encoded gene NTH1 and the proline oxidase-encoded gene PUT1. We first used the two-step integration-based seamless gene deletion method to separately delete NTH1 and PUT1 in haploid yeast. Subsequently, through two rounds of hybridization and sporulation-based allelic exchange and colony PCR-mediated tetrad analysis, we obtained strains with restored URA3 and deletion of NTH1 and/or PUT1. The resulting strain showed higher cell survival and dough-leavening ability after freezing compared to the wild-type strain due to enhanced accumulation of trehalose and/or proline. Moreover, mutant with simultaneous deletion of NTH1 and PUT1 exhibits the highest relative dough-leavening ability after freezing compared to mutants with single-gene deletion perhaps due to elevated levels of both trehalose and proline. These results verified that it is applicable to construct frozen dough baker's yeast using the method proposed in this paper. SN - 1476-5535 UR - https://www.unboundmedicine.com/medline/citation/26965428/Improving_freeze_tolerance_of_baker's_yeast_through_seamless_gene_deletion_of_NTH1_and_PUT1_ L2 - http://dx.doi.org/10.1007/s10295-016-1753-7 DB - PRIME DP - Unbound Medicine ER -