Abstract
Expression and purification of recombinant proteins produced in plants is emerging as an affordable alternative to using more costly mammalian bioreactors since plants are capable of producing mammalian proteins at high concentrations. There are two general methods of expressing foreign proteins in plants, namely, transient expression and stable transgenic expression. Both methods have advantages which serve different purposes. Nicotiana benthamiana is primarily used as plant host for transient expression of foreign proteins. This system is capable of producing high yields of antibody in a relatively short period of time (days); however, intensive upstream processing is required as each plant must be infected with Agrobacterium tumefaciens cells by vacuum infiltration. N. tabacum is often used for production of stable transgenic plants through a procedure that requires longer development time (months). Although antibody yields are smaller compared with the transient method, the advantage of using stable transgenic expression is that very little upstream process management is required once homozygous seed lines are developed. In this chapter, we describe the basic methodologies for expressing antibodies in plants using the transient and transgenic systems.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Gelvin SB (2003) Agrobacterium-mediated plant transformation: the biology behind the “gene-jockeying” tool. Microbiol Mol Biol Rev 67(1):16–37
Lee LY, Gelvin SB (2008) T-DNA binary vectors and systems. Plant Physiol 146(2):325–332
Veluthambi K, Jayaswal RK, Gelvin SB (1987) Virulence genes A, G, and D mediate the double-stranded border cleavage of T-DNA from the Agrobacterium Ti plasmid. Proc Natl Acad Sci U S A 84(7):1881–1885
Gallois P, Marinho P (1995) Leaf disk transformation using Agrobacterium tumefaciens-expression of heterologous genes in tobacco. Methods Mol Biol 49:39–48
Gleba Y, Klimyuk V, Marillonnet S (2005) Magnifection—a new platform for expressing recombinant vaccines in plants. Vaccine 23(17–18):2042–2048
Gleba Y, Klimyuk V, Marillonnet S (2007) Viral vectors for the expression of proteins in plants. Curr Opin Biotechnol 18(2):134–141
Marillonnet S, Giritch A, Gils M et al (2004) In planta engineering of viral RNA replicons: efficient assembly by recombination of DNA modules delivered by Agrobacterium. Proc Natl Acad Sci U S A 101(18):6853–6857
Chebolu S, Daniell H (2009) Chloroplast-derived vaccine antigens and biopharmaceuticals: expression, folding, assembly and functionality. Curr Top Microbiol Immunol 332:33–54
Rasala BA, Muto M, Lee PA et al (2010) Production of therapeutic proteins in algae, analysis of expression of seven human proteins in the chloroplast of Chlamydomonas reinhardtii. Plant Biotechnol J 8(6):719–733
Halpin C (2005) Gene stacking in transgenic plants—the challenge for 21st century plant biotechnology. Plant Biotechnol J 3(2):141–155
Castilho A, Strasser R, Stadlmann J et al (2010) In planta protein sialylation through overexpression of the respective mammalian pathway. J Biol Chem 285(21):15923–15930
Cox KM, Sterling JD, Regan JT et al (2006) Glycan optimization of a human monoclonal antibody in the aquatic plant Lemna minor. Nat Biotechnol 24(12):1591–1597
Outchkourov NS, Peters J, de Jong J et al (2003) The promoter-terminator of chrysanthemum rbcS1 directs very high expression levels in plants. Planta 216(6):1003–1012
De Muynck B, Navarre C, Boutry M (2010) Production of antibodies in plants: status after twenty years. Plant Biotechnol J 8(5):529–563
Odell JT, Nagy F, Chua NH (1985) Identification of DNA sequences required for activity of the cauliflower mosaic virus 35S promoter. Nature 313(6005):810–812
Lee LY, Kononov ME, Bassuner B et al (2007) Novel plant transformation vectors containing the superpromoter. Plant Physiol 145(4):1294–1300
Zaidi MA, Ye G, Yao H et al (2009) Transgenic rice plants expressing a modified cry1Ca1 gene are resistant to Spodoptera litura and Chilo suppressalis. Mol Biotechnol 43(3):232–242
Grohs BM, Niu Y, Veldhuis LJ et al (2010) Plant-produced trastuzumab inhibits the growth of HER2 positive cancer cells. J Agric Food Chem 58(18):10056–10063
Cheung SC, Sun SS, Chan JC et al (2009) Expression and subcellular targeting of human insulin-like growth factor binding protein-3 in transgenic tobacco plants. Transgenic Res 18(6):943–951
Lingner T, Kataya AR, Antonicelli GE et al (2011) Identification of novel plant peroxisomal targeting signals by a combination of machine learning methods and in vivo subcellular targeting analyses. Plant Cell 23(4):1556–1572
Sugio T, Satoh J, Matsuura H et al (2008) The 5′-untranslated region of the Oryza sativa alcohol dehydrogenase gene functions as a translational enhancer in monocotyledonous plant cells. J Biosci Bioeng 105(3):300–302
Butaye KM, Goderis IJ, Wouters PF et al (2004) Stable high-level transgene expression in Arabidopsis thaliana using gene silencing mutants and matrix attachment regions. Plant J 39(3):440–449
Giritch A, Marillonnet S, Engler C et al (2006) Rapid high-yield expression of full-size IgG antibodies in plants coinfected with noncompeting viral vectors. Proc Natl Acad Sci U S A 103(40):14701–14706
Marillonnet S, Thoeringer C, Kandzia R et al (2005) Systemic Agrobacterium tumefaciens-mediated transfection of viral replicons for efficient transient expression in plants. Nat Biotechnol 23(6):718–723
Mandahar CL (2006) Multiplication of RNA plant viruses. Springer, Dordrecht
Meyers AJ, Grohs BM, Hall JC (2011) Antibody production in planta. In: Butler M, Webb C, Moreira A, Grodzinski B, Cui ZF, Moo-Young M (eds) Comprehensive biotechnology, 2nd edn. Elsevier, Oxford
McLean MD, Almquist KC, Niu Y et al (2007) A human anti-Pseudomonas aeruginosa serotype O6ad immunoglobulin G1 expressed in transgenic tobacco is capable of recruiting immune system effector function in vitro. Antimicrob Agents Chemother 51(9):3322–3328
De Muynck B, Navarre C, Boutry M (2010) Production of antibodies in plants: status after twenty years. Plant Biotechnol J 8(5):529–563
Conley AJ, Zhu H, Le LC et al (2011) Recombinant protein production in a variety of Nicotiana hosts: a comparative analysis. Plant Biotechnol J 9(4):434–444
Almquist KC, McLean MD, Niu Y et al (2006) Expression of an anti-botulinum toxin A neutralizing single-chain Fv recombinant antibody in transgenic tobacco. Vaccine 24(12):2079–2086
Makvandi-Nejad S, McLean MD, Hirama T et al (2005) Transgenic tobacco plants expressing a dimeric single-chain variable fragment (scfv) antibody against Salmonella enterica serotype Paratyphi B. Transgenic Res 14(5):785–792
Olea-Popelka F, McLean MD, Horsman J et al (2005) Increasing expression of an anti-picloram single-chain variable fragment (ScFv) antibody and resistance to picloram in transgenic tobacco (Nicotiana tabacum). J Agric Food Chem 53(17):6683–6690
Acknowledgements
Funding was provided to J.C.H. from the Ontario Ministry of Agriculture, Food and Rural Affairs (OMAFRA), the Canada Research Chairs (CRC) program of the Natural Sciences and Engineering Council of Canada (NSERC), the Ontario Centres of Excellence (OCE) program of the Ontario Ministry of Research and Innovation (OMRI), and PlantForm Corporation. Magnifection vectors were provided by Icon Genetics, GmbH.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2012 Springer Science+Business Media, LLC
About this protocol
Cite this protocol
Garabagi, F., McLean, M.D., Hall, J.C. (2012). Transient and Stable Expression of Antibodies in Nicotiana Species. In: Chames, P. (eds) Antibody Engineering. Methods in Molecular Biology, vol 907. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-61779-974-7_23
Download citation
DOI: https://doi.org/10.1007/978-1-61779-974-7_23
Published:
Publisher Name: Humana Press, Totowa, NJ
Print ISBN: 978-1-61779-973-0
Online ISBN: 978-1-61779-974-7
eBook Packages: Springer Protocols