Screening efficient constitutive promoters in Corynebacterium glutamicum based on time-series transcriptome analysis.

Yingchun Wang; Jiao Liu; Xiaomeng NI; Yu Lei; Ping Zheng; Aipo Diao

Return

Screening efficient constitutive promoters in Corynebacterium glutamicum based on time-series transcriptome analysis.

Author: Yingchun WANG ¹ ; Jiao LIU ² ; Xiaomeng NI ² ; Yu LEI ³ ; Ping ZHENG ² ; Aipo DIAO ¹
Author Information

1. School of Biological Engineering, Tianjin University of Science and Technology, Tianjin 300457, China.
2. Key Laboratory of Systems Microbial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China.
3. Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China.
Publication Type:Journal Article
Keywords: Corynebacterium glutamicum; promoter; red fluorescent protein reporter system; time-series transcriptome
MeSH: Corynebacterium glutamicum; genetics; Gene Expression Profiling; Gene Expression Regulation, Bacterial; Metabolic Engineering; Promoter Regions, Genetic; Transcriptome
From: Chinese Journal of Biotechnology 2018;34(11):1760-1771
CountryChina
Language:Chinese
Abstract: Promoter, an essential regulatory element, is widely used for metabolic engineering of industrial strains. Corynebacterium glutamicum is an important industrial workhorse to produce various amino acids. However, strong constitutive promoters that are applicable to C. glutamicum are rarely reported. In this study, we first performed a time-series transcriptome analysis of a glutamate hyper-producing strain C. glutamicum SL4 by using RNA-Seq. Overall, we picked 10 samples at different time during the fermentation process. By analyzing the time-series transcriptome data, we selected 10 candidate genes with the highest transcriptional level. These genes were all transcribed stably during the fermentation process. We subsequently cloned the promoter sequences and evaluated the promoters' strength in strain SL4 using a red fluorescent protein reporter system. To evaluate the universality of the promoters in different C. glutamicum strains, we further tested the performance of some promoters in wild type C. glutamicum strains, including ATCC 13869 and ATCC 13032. The strongest promoter was further characterized using LacZ as a reporter in all the three C. glutamicum strains. Finally, we successfully obtained three constitutive promoters with universality, PcysK, PgapA and PfumC. PcysK is the most efficient promoter among the three C. glutamicum strains. In strains SL4 and ATCC 13869, the strength of PcysK is 2-fold of the strong inducible promoter Ptac using the red fluorescent protein as a reporter and 4-fold of Ptac using LacZ as a reporter. Moreover, the strength of PcysK reaches 30%-40% of Ptac in strain ATCC 13032. The promoter PcysK is identified as a strong promoter for the first time, which can be used as an efficient biobrick for metabolic engineering of synthesis pathways in C. glutamicum.