title: Isolation of strong constitutive promoters from Lactococcus lactis subsp. lactis N8.
abstract: The synthesis of heterologous proteins in Lactococcus lactis is strongly influenced by the promoter selected for the expression. The nisin A promoter is commonly used for induced expression of proteins in L. lactis, whereas few constitutive promoters (P45 and the weaker P32) have been used for protein expression studies. In this study, eight different putative strong constitutive promoters were identified through transcriptional analysis of L. lactis N8 and were investigated for their capability to drive nisZ gene expression with promoters P45 and P32 as control. Four strong promoters (P8, P5, P3 and P2) were identified as having a transcriptional activity that was higher than that of P45 through RT-qPCR and agar-diffusion experiments. In addition, these four promoters were fused to the erythromycin resistant gene (ermC) with promoter P45 as control and inserted into the backbone of the pNZ8048 vector. The transcriptional efficiencies of promoters P8, P5, P2 and P3 were all higher than promoter P45 based on the obtained MIC50 values and they all showed different activity levels. In conclusion, four strong constitutive promoters with a wide range of promoter activities were identified and are suitable for protein production in L. lactis.
authors.affiliation: College of Life Sciences, The Key Laboratory of Molecular Microbiology and Technology, Ministry of Education, Nankai University, Tianjin, China.
authors.forename: Duolong
authors.initials: D
authors.lastname: Zhu
authors.affiliation: College of Life Sciences, The Key Laboratory of Molecular Microbiology and Technology, Ministry of Education, Nankai University, Tianjin, China.
authors.forename: Fulu
authors.initials: F
authors.lastname: Liu
authors.affiliation: College of Life Sciences, The Key Laboratory of Molecular Microbiology and Technology, Ministry of Education, Nankai University, Tianjin, China.
authors.forename: Haijin
authors.initials: H
authors.lastname: Xu
authors.affiliation: College of Life Sciences, The Key Laboratory of Molecular Microbiology and Technology, Ministry of Education, Nankai University, Tianjin, China.
authors.forename: Yanling
authors.initials: Y
authors.lastname: Bai
authors.affiliation: College of Life Sciences, The Key Laboratory of Molecular Microbiology and Technology, Ministry of Education, Nankai University, Tianjin, China.
authors.forename: Xiuming
authors.initials: X
authors.lastname: Zhang
authors.affiliation: Department of Food and Environmental Sciences, University of Helsinki, Finland.
authors.forename: Per Erik Joakim
authors.initials: PE
authors.lastname: Saris
authors.affiliation: College of Life Sciences, The Key Laboratory of Molecular Microbiology and Technology, Ministry of Education, Nankai University, Tianjin, China mingqiangqiao@aliyun.com.
authors.forename: Mingqiang
authors.initials: M
authors.lastname: Qiao
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics*; D001483:Base Sequence; D015964:Gene Expression Regulation, Bacterial; D005822:Genetic Vectors; D013294:Lactococcus lactis / Q000235:genetics*; D009561:Nisin / Q000235:genetics; D011401:Promoter Regions, Genetic; D011499:Protein Processing, Post-Translational; D014158:Transcription, Genetic
chemical_list: D001426:Bacterial Proteins; D009561:Nisin; C102526:nisin A
keywords: Lactococcus lactis; MIC50; nisZ; strong promoters; transcriptional analysis