Efficient agroinfiltration of plants for high-level transient expression of recombinant proteins

Kahlin Leuzinger, Matthew Dent, Jonathan Hurtado, Jake Stahnke, Huafang Lai, Xiaohong Zhou, Qiang Chen

Research output: Contribution to journalArticlepeer-review

90 Scopus citations

Abstract

Mammalian cell culture is the major platform for commercial production of human vaccines and therapeutic proteins. However, it cannot meet the increasing worldwide demand for pharmaceuticals due to its limited scalability and high cost. Plants have shown to be one of the most promising alternative pharmaceutical production platforms that are robust, scalable, low-cost and safe. The recent development of virus-based vectors has allowed rapid and high-level transient expression of recombinant proteins in plants. To further optimize the utility of the transient expression system, we demonstrate a simple, efficient and scalable methodology to introduce target-gene containing Agrobacterium into plant tissue in this study. Our results indicate that agroinfiltration with both syringe and vacuum methods have resulted in the efficient introduction of Agrobacterium into leaves and robust production of two fluorescent proteins; GFP and DsRed. Furthermore, we demonstrate the unique advantages offered by both methods. Syringe infiltration is simple and does not need expensive equipment. It also allows the flexibility to either infiltrate the entire leave with one target gene, or to introduce genes of multiple targets on one leaf. Thus, it can be used for laboratory scale expression of recombinant proteins as well as for comparing different proteins or vectors for yield or expression kinetics. The simplicity of syringe infiltration also suggests its utility in high school and college education for the subject of biotechnology. In contrast, vacuum infiltration is more robust and can be scaled-up for commercial manufacture of pharmaceutical proteins. It also offers the advantage of being able to agroinfiltrate plant species that are not amenable for syringe infiltration such as lettuce and Arabidopsis. Overall, the combination of syringe and vacuum agroinfiltration provides researchers and educators a simple, efficient, and robust methodology for transient protein expression. It will greatly facilitate the development of pharmaceutical proteins and promote science education.

Original languageEnglish (US)
Pages (from-to)e50521
JournalJournal of Visualized Experiments
Issue number77
DOIs
StatePublished - Jul 23 2013

Keywords

  • Agrobacterium tumefaciens
  • Agroinfiltration
  • Antibodies
  • Bioengineering
  • Cellular biology
  • DsRed
  • GFP
  • Geminiviral vectors
  • Gene expression
  • Gene transfer techniques
  • Genetics
  • Green fluorescent proteins
  • Imaging
  • Issue 77
  • Microbiology
  • Molecular biology
  • Monoclonal
  • Monoclonal antibody
  • Nicotiana benthamiana
  • Plant biology
  • Plant infiltration
  • Plant model
  • Plant proteins
  • Plant viruses
  • Plant-made pharmaceuticals
  • Recombinant proteins
  • Synthetic
  • Syringe agroinfiltration
  • Vaccines
  • Vacuum agroinfiltration
  • Virology
  • Virus-like particle

ASJC Scopus subject areas

  • Neuroscience(all)
  • Chemical Engineering(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Immunology and Microbiology(all)

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