Construction of expression vector and selection of promoter in Lactococcus lactis

• 1、School of pharmcy,Nanjing University of Chinese Medicine,Nanjing Jiangsu,210023
• 2、Nanjing Genrecom biotechnology Co,LTD,Nanjing,Jiangsu 210019
• 3、School of life Sciences,Nanjing Unversity,Nanjing,Jiangsu 210023
• 4、Institute of Pharmaceutical Biotechnology,Jiangsu Institute of industrial Technology，Nanjing,Jiangsu 210042

Abstract：Lactococcus lactis is a kind of probiotics that is abundant in the intestinal tract. Its metabolism is relatively simple, the intracellular and extracellular proteins are easy to be separated, and it does not produce endotoxins. It is an ideal gene expression host bacteria that can be used in the human body. In order to screen the promoters suitable for genetic engineering expression in L. lactis, two recombinant L. lactis expression plasmids pLCpPRPL-RFP and pLCpnis-RFP were constructed in this study. The plasmid pLCpPRPL-RFP employs the heat shock promoter pPRPL, and the pLCpnis-RFP plasmid uses the inducible promoter pnis by Nisin. Both expression plasmids use the same RFP (red fluorescent protein) as the reporter. After optimizing the conditions of electro-transformation, the two expression plasmids were electro-transformed into Lactococcus lactis NZ9000, and the recombinant Lactococcus lactis NZ9000 was induced to express the red fluorescent protein RFP. By comparing the fluorescence absorption values of two recombinant expression vectors expressing red fluorescent protein, it is proved that the temperature-controlled promoter pPRPL has stronger expression efficiency, and the cost of inducing expression is lower. This study made a useful exploration for the construction and application of genetically engineered Lactococcus lactis.

Keywords： Genetic engineering Lactococcus lactis Promoter Red fluorescent protein