Plasmid pVAX1-NH36 purification by membrane and bead perfusion chromatography

Diana Ivonne Franco-Medrano, Patricia Guerrero-Germán*, Rosa María Montesinos-Cisneros, Jaime Ortega-López, Armando Tejeda-Mansir

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

7 Scopus citations

Abstract

The demand for plasmid DNA (pDNA) has increased in response to the rapid advances in vaccines applications to prevent and treat infectious diseases caused by virus, bacteria or parasites, such as Leishmania species. The immunization protocols require large amounts of supercoiled plasmid DNA (sc-pDNA) challenging the development of efficient and profitable processes for capturing and purified pDNA molecules from large volumes of lysates. A typical bioprocess involves four steps: fermentation, primary recovery, intermediate recovery and final purification. Ion-exchange chromatography is one of the key operations in the purification schemes of pDNA owing the chemical structure of these macromolecules. The goal of this research was to compare the performance of the final purification step of pDNA using ion-exchange chromatography on columns packed with Mustang Q membranes or perfusive beads POROS 50 HQ. The experimental results showed that both matrixes could separate the plasmid pVAX1-NH36 (3936 bp) from impurities in clarified Escherichia coli lysates with an adequate resolution. In addition, a 24- and 21-fold global purification factor was obtained. An 88 and 63% plasmid recuperation was achieved with ion-exchange membranes and perfusion beads, respectively. A better understanding of perfusion-based matrices for the purification of pDNA was developed in this research.

Original languageEnglish
Pages (from-to)463-471
Number of pages9
JournalBioprocess and Biosystems Engineering
Volume40
Issue number3
DOIs
StatePublished - Mar 2017

Bibliographical note

Publisher Copyright:
© 2016, Springer-Verlag Berlin Heidelberg.

Keywords

  • Leishmaniasis
  • Perfusion chromatography
  • Plasmid DNA
  • Vaccines

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