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Temporary Immersion Bioreactors for the Contained Production of Recombinant Proteins in Transplastomic Plants

  • Sherwin Barretto
  • Franck MichouxEmail author
  • Peter J. Nixon
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1385)

Abstract

Despite the largely maternal inheritance of plastid genomes, the risk of transgene dissemination from transplastomic plants can limit the scope for field cultivation. There is a need for a cost-effective, scalable process to grow large quantities of transplastomic plant biomass for biosynthesis of biopharmaceuticals and other high-value heterologous proteins. Temporary immersion culture is a means of achieving this under fully contained conditions. This method describes the organogenesis of transplastomic Nicotiana tabacum callus in RITA® temporary immersion bioreactors to produce rootless leafy biomass, and subsequent total soluble protein extraction, SDS-PAGE, and Western immunoblot analysis of heterologous protein expression. This method can be used for propagation of plastid or nuclear transformants, though is especially suitable for transplastomic biomass, as organogenesis leads to greater expression and accumulation of transplastomic proteins due to increases in chloroplast number and size.

Keywords

Temporary immersion bioreactors Transgene containment Transplastomic Heterologous protein expression Micropropagation Morphogenesis 

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Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  • Sherwin Barretto
    • 1
  • Franck Michoux
    • 2
    Email author
  • Peter J. Nixon
    • 1
  1. 1.Sir Ernst Chain Building-Wolfson Laboratories, Department of LifeImperial College LondonLondonUK
  2. 2.Alkion BiopharmaEvryFrance

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