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Plasmid pVAX1-NH36 purification by membrane and bead perfusion chromatography

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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.

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Acknowledgements

This research was supported by the grant provided by the National Council for Science and Technology (CONACYT) under the projects CB2012-179779 and CB2015-257411. We also appreciate the support given by the Strengthening Program Quality in Educative Institutions (PROFOCIE) 2015 and the University of Sonora.

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Authors and Affiliations

  1. Departamento de Ingeniería Química y Metalurgia, Universidad de Sonora, Blvd. Luis Encinas y Rosales s/n, 83000, Hermosillo, SON, México

    Diana Ivonne Franco-Medrano & Patricia Guerrero-Germán

  2. Departamento de Matemáticas, Universidad de Sonora, Hermosillo, SON, México

    Rosa María Montesinos-Cisneros

  3. Departamento de Biotecnología y Bioingeniería, Centro de Investigación y de Estudios Avanzados del Instituto Politécnico NacionalAv. Instituto Politécnico Nacional # 2508, Cd. De, 07360, México, México

    Jaime Ortega-López

  4. Departamento de Investigaciones Científicas y Tecnológicas, Universidad de Sonora, Blvd. Luis Encinas s/n, 83000, Hermosillo, SON, México

    Armando Tejeda-Mansir

Authors
  1. Diana Ivonne Franco-Medrano
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  2. Patricia Guerrero-Germán
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  3. Rosa María Montesinos-Cisneros
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  4. Jaime Ortega-López
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  5. Armando Tejeda-Mansir
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Correspondence to Patricia Guerrero-Germán.

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Franco-Medrano, D.I., Guerrero-Germán, P., Montesinos-Cisneros, R.M. et al. Plasmid pVAX1-NH36 purification by membrane and bead perfusion chromatography. Bioprocess Biosyst Eng 40, 463–471 (2017). https://doi.org/10.1007/s00449-016-1714-6

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  • DOI: https://doi.org/10.1007/s00449-016-1714-6

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