Abstract
This chapter constitutes a practical guide to using the “pEAQ” vector series for transient or stable expression of one or more protein(s) in Nicotiana benthamiana plants. The pEAQ vectors are a series of small binary vectors designed for controlled expression of multiple proteins in plants. To achieve high levels of expression, an expression system based on translational enhancement by the untranslated regions of RNA-2 from cowpea mosaic virus (CPMV), named CPMV-HT, is used. The expression vector pEAQ-HT combines the user-friendly pEAQ plasmid with CPMV-HT to provide a system for high-level expression of proteins in plants.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Giritch A, Marillonnet S, Engler C, van Eldik G, Botterman J, Klimyuk V, Gleba Y (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
Sainsbury F, Lavoie PO, D'Aoust MA, Vezina LP, Lomonossoff GP (2008) Expression of multiple proteins using full-length and deleted versions of cowpea mosaic virus RNA-2. Plant Biotechnol J 6(1):82–92
Sainsbury F, Lomonossoff GP (2008) Extremely high-level and rapid transient protein production in plants without the use of viral replication. Plant Physiol 148(3):1212–1218
Sainsbury F, Thuenemann EC, Lomonossoff GP (2009) pEAQ: versatile expression vectors for easy and quick transient expression of heterologous proteins in plants. Plant Biotechnol J 7(7):682–693
Voinnet O, Rivas S, Mestre P, Baulcombe D (2003) An enhanced transient expression system in plants based on suppression of gene silencing by the p19 protein of tomato bushy stunt virus. Plant J 33(5):949–956
Sainsbury F, Saxena P, Geisler K, Osbourn A, Lomonossoff GP (2012) Using a virus-derived system to manipulate plant natural product biosynthetic pathways. Methods Enzymol 517:185–202
Thuenemann EC, Lenzi P, Love AJ, Taliansky M, Bécares M, Zuñiga S, Enjuanes L, Zahmanova GG, Minkov IN, Matić S, Noris E, Meyers A, Hattingh A, Rybicki EP, Kiselev OI, Ravin NV, Eldarov MA, Skriabin KG, Lomonossoff GP (2013) The use of transient expression systems for the rapid production of virus-like particles in plants. Curr Pharm Des 19(31):5564–5573
Vardakou M, Sainsbury F, Rigby N, Mulholland F, Lomonossoff GP (2012) Expression of active recombinant human gastric lipase in Nicotiana benthamiana using the CPMV-HT transient expression system. Protein Expr Purif 81(1):69–74
Sainsbury F, Sack M, Stadlmann J, Quendler H, Fischer R, Lomonossoff GP (2010) Rapid transient production in plants by replicating and non-replicating vectors yields high quality functional anti-HIV antibody. PLoS One 5(11):e13976
Thuenemann EC, Meyers AE, Verwey J, Rybicki EP, Lomonossoff GP (2013) A method for rapid production of heteromultimeric protein complexes in plants: assembly of protective bluetongue virus-like particles. Plant Biotechnol J 11(7):839–846
Matic S, Rinaldi R, Masenga V, Noris E (2011) Efficient production of chimeric human papillomavirus 16 L1 protein bearing the M2e influenza epitope in Nicotiana benthamiana plants. BMC Biotechnol 11:106
Peyret H, Lomonossoff GP (2013) The pEAQ vector series: the easy and quick way to produce recombinant proteins in plants. Plant Mol Biol 83(1–2):51–58
Montague NP, Thuenemann EC, Saxena P, Saunders K, Lenzi P, Lomonossoff GP (2011) Recent advances of cowpea mosaic virus-based particle technology. Hum Vaccin 7(3):|383–390
Hoekema A, Hirsch PR, Hooykaas PJJ, Schilperoort RA (1983) Binary vector strategy based on separation of vir- and T-region of the Agrobacterium tumefaciens Ti-plasmid. Nature 303:179–180
Meshcheriakova YA, Saxena P, Lomonossoff GP (2014) Fine-tuning levels of heterologous gene expression in plants by orthogonal variation of the untranslated regions of a nonreplicating transient expression system. Plant Biotechnol J 12(6):718–727
Saxena P, Hsieh YC, Alvarado VY, Sainsbury F, Saunders K, Lomonossoff GP, Scholthof HB (2011) Improved foreign gene expression in plants using a virus-encoded suppressor of RNA silencing modified to be developmentally harmless. Plant Biotechnol J 9(6):703–712
Acknowledgement
This work was supported by grant BB/J004561/1 from the Biotechnology and Biological Research Council (BBSRC) UK and the John Innes Foundation.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2016 Springer Science+Business Media New York
About this protocol
Cite this protocol
Saxena, P., Thuenemann, E.C., Sainsbury, F., Lomonossoff, G.P. (2016). Virus-Derived Vectors for the Expression of Multiple Proteins in Plants. In: MacDonald, J., Kolotilin, I., Menassa, R. (eds) Recombinant Proteins from Plants. Methods in Molecular Biology, vol 1385. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3289-4_3
Download citation
DOI: https://doi.org/10.1007/978-1-4939-3289-4_3
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-3288-7
Online ISBN: 978-1-4939-3289-4
eBook Packages: Springer Protocols