title: Increasing plasmid-based DNA vaccine construct (16 kb pSVK-HBVA) production inEscherichia coliXL10-Gold through optimization of media component.
abstract: At present, there are production processes to produce protein byEscherichia coli(E. coli) fermentation. Research on the design and optimization of the plasmid fermentation medium, however, is less advanced. The fermentation medium that is optimized for plasmid DNA production is different from the medium that is optimized for protein production. So, establishing a scientific and rational method to optimize the fermentation medium used for plasmid production is very important. Previously, our laboratory developed a novel therapeutic DNA vaccine (named pSVK-HBVA) for hepatitis B based on the alphavirus replicon, and found thatE. coliXL10-Gold was the optimal host strain for the production of plasmid pSVK-HBVA. The aim of this study was to establish a scientific and rational method to optimize the fermentation medium used for plasmid production, and investigate the effect of growth medium composition on the production of plasmid pSVK-HBVA harboured inE. coliXL10-Gold, as well as to optimize the medium composition. The one-factor-at-a-time experiments demonstrated that Luria-Bertani (LB) was the optimal basic medium. The optimal carbon source and nitrogen source were glycerol and home-made proteose peptone, respectively. Based on the Plackett-Burman (PB) design, proteose peptone, glycerol and NH4Cl were identified as the significant variables, which were further optimized by the steepest ascent (descent) method and central composite design. Growth medium optimization in 500-mL shake flasks by response surface methodology resulted in a maximum volumetric yield of 13.61 mg/L, which was approximately 2.5 times higher than that obtained from the basic medium (LB).
authors.affiliation: Institute of Basic Medical Science, Academy of Military Medical Sciences , 27 Tai Ping Road, Beijing 100850 , China.
authors.forename: Yu
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Institute of Basic Medical Science, Academy of Military Medical Sciences , 27 Tai Ping Road, Beijing 100850 , China.
authors.forename: Liang
authors.initials: L
authors.lastname: Zhang
authors.affiliation: Institute of Basic Medical Science, Academy of Military Medical Sciences , 27 Tai Ping Road, Beijing 100850 , China.
authors.forename: Wei
authors.initials: W
authors.lastname: Zhang
authors.affiliation: Institute of Basic Medical Science, Academy of Military Medical Sciences , 27 Tai Ping Road, Beijing 100850 , China.
authors.forename: Hao
authors.initials: H
authors.lastname: Wu
authors.affiliation: Institute of Basic Medical Science, Academy of Military Medical Sciences , 27 Tai Ping Road, Beijing 100850 , China.
authors.forename: Xiao Ming
authors.initials: XM
authors.lastname: Zhu
authors.affiliation: Institute of Basic Medical Science, Academy of Military Medical Sciences , 27 Tai Ping Road, Beijing 100850 , China.
authors.forename: Yuan Ji
authors.initials: YJ
authors.lastname: Xu
authors.affiliation: Institute of Basic Medical Science, Academy of Military Medical Sciences , 27 Tai Ping Road, Beijing 100850 , China.
authors.forename: Jin Qi
authors.initials: JQ
authors.lastname: Yan
authors.affiliation: Institute of Basic Medical Science, Academy of Military Medical Sciences , 27 Tai Ping Road, Beijing 100850 , China.
authors.forename: Ji Yun
authors.initials: JY
authors.lastname: Yu
keywords: DNA vaccine; Escherichia coli; fermentation media; media component optimization