Industrial scale production of plasmid DNA for vaccine and gene therapy: plasmid design, production, and purification

  title={Industrial scale production of plasmid DNA for vaccine and gene therapy: plasmid design, production, and purification},
  author={Kristala Jones Prather and Sangeetha L. Sagar and Jason C. Murphy and Michel Chartrain},
  journal={Enzyme and Microbial Technology},

Figures and Tables from this paper

Upstream processing of plasmid DNA for vaccine and gene therapy applications.
This review summarizes recent patents and patent applications relating to plasmid upstream processing manufacturing, ranging fromplasmid design to growth strategies to produce plasmID-bearing E. coli.
Comparative analysis of enzymatically produced novel linear DNA constructs with plasmids for use as DNA vaccines
This work describes a novel, double-stranded, linear DNA construct produced by an enzymatic process that solely encodes an antigen expression cassette, comprising antigen, promoter, polyA tail and telomeric ends, and demonstrates that expression levels were equivalent between Doggybones and plasmids both in vitro and in vivo.
Development of a Highly Productive and Scalable Plasmid DNA Production Platform
This second generation process, based on a defined cultivation medium and feed solution excluding MSG, proved to be scalable, robust, and highly productive.
Industrial Manufacturing of Plasmid-DNA Products for Gene Vaccination and Therapy
The state of the art in industrial plasmid manufacturing is summarized and bottlenecks and pitfalls to be avoided are discussed in order to obtain pDNA products of high and reproducible quantity and quality.
Factors influencing the successful production of large plasmids for use in gene therapy and DNA vaccination.
Investigations conducted to determine issues which may be faced should attempts be made to produce large plasmids (>3-10kb) for use in areas such as DNA vaccination and gene therapy and the binding behaviour of the large plasmsids to chromatographic resins used for plasmid purification are outlined.
Plasmid DNA production for therapeutic applications.
In this chapter, the different stages of plasmid DNA production are reviewed, from the vector design to downstream operation options, and recent advances on cell engineering for improving plasmids production are discussed.
Scalable Technology to Produce Pharmaceutical Grade Plasmid DNA for Gene Therapy
The use of gene therapy is a promising process for the prevention, treatment and cure of diseases such as cancer and acquired inmudeficiency syndrome (AIDS); increasing a considerable interest during
Improved downstream process for the production of plasmid DNA for gene therapy.
A novel economic downstream process which overcomes the bottlenecks of common lab-scale techniques and meets all regulatory requirements to satisfy the increasing demand for pharmaceutical grade plasmids.
Using Plasmids as DNA Vaccines for Infectious Diseases.
This review will introduce some key concepts in the use of DNA plasmids as vaccines, including how the DNA enters the cell and is expressed, how it induces an immune response, and a summary of clinical trials with DNA vaccines.
Development of new plasmid DNA vaccine vectors with R 1-based replicons Citation
It is demonstrated that R1-based plasmids can produce high yields of high-quality pDNA without the need for a temperature shift, and the groundwork for further investigation of this class of vectors in the context of plasmid DNA production is laid.


Cancer gene therapy using plasmid DNA: purification of DNA for human clinical trials.
The scaleable purification method described here is a combination of highly reproducible unit operations; alkaline lysis, precipitation, and size-exclusion chromatography, and the advantages over existing DNA purification methods include improved plasmid purity and the elimination of undesirable process additives such as toxic organic extractants and animal-derived enzymes.
Depression of protein synthetic capacity due to cloned‐gene expression in E. coli
The data indicate that a limitation develops within the translational capacity of the cell at high levels of cloned‐gene expression and suggest that strategies designed to enhance recombinant protein expression should include manipulation of translation as well as transcription.
High-yield production of pBR322-derived plasmids intended for human gene therapy by employing a temperature-controllable point mutation.
The plasmid DNA isolated from this process contained lower levels of RNA and chromosomal DNA contaminants, simplifying downstream processing.
DNA and RNA-based vaccines: principles, progress and prospects.
Purification of a cystic fibrosis plasmid vector for gene therapy using hydrophobic interaction chromatography
A new method is described for the purification of a cystic fibrosis plasmid vector of clinical grade, which includes an ammonium sulfate precipitation followed by hydrophobic interaction chromatography (HIC) using a Sepharose gel derivatized with 1,4‐butanediol‐diglycidylether.
Genetic immunization: a new era in vaccines and immune therapeutics
It is hoped that DNA inoculation will ultimately lead to new vaccines that are immunologically effective and economically accessible to all nations.
High copy number plasmids compatible with commonly used cloning vectors.
To maximize expression of two different gene products, the aim was to construct new cloning vectors that are compatible with many commonly used plasmids and that replicate at a high copy number.
Repressor titration: a novel system for selection and stable maintenance of recombinant plasmids.
A novel system that employs plasmid-mediated repressor titration to activate a chromosomal selectable marker, removing the requirement for a plasid-borne marker gene is described.