Antibody-Mediated Pathogen Resistance in Plants
The methods described in this chapter were developed in order to produce transgenic plants expressing pathogen-specific single-chain variable fragment (scFv) antibodies fused to antifungal peptides (AFPs), conferring resistance against fungal pathogens. We describe the selection from a phage display library of avian scFv antibodies that recognize cell surface proteins on fungi from the genus Fusarium, and the construction of scFv–AFP fusion protein constructs followed by their transient expression in tobacco (Nicotiana spp.) plants and stable expression in Arabidopsis thaliana plants. Using these techniques, the antibody fusion with the most promising in vitro activity can be used to generate transgenic plants that are resistant to pathogens such as Fusarium oxysporum f. sp. matthiolae.
KeywordsTransgenic plants Antibody Antibody fragments Single-chain variable fragment scFv Pathogen Fungal antigens Antifungal peptide Arabidopsis thaliana Fusarium spp.
We would like to thank all our colleagues at the Fraunhofer IME and the Institute of Biology VII, RWTH Aachen, for their cooperation and helpful discussions. Special thanks to Drs. Yu-Cai Liao and Simone Dorfmüller for their collaboration with the Fusarium work. We would like to thank Dr. Richard M. Twyman for help with the preparation of this manuscript.
- 10.Nölke G, Fischer R, Schillberg S (2004) Antibody-based pathogen resistance in plants. J Plant Pathol 86:5–17Google Scholar
- 24.Cheng W, Li HP, Zhang JB, Du HJ, Wei QY, Huang T, Yang P, Kong XW, Liao YC (2015) Tissue-specific and pathogen-inducible expression of a fusion protein containing a Fusarium-specific antibody and a fungal chitinase protects wheat against Fusarium pathogens and mycotoxins. Plant Biotechnol J 13(5):664–674CrossRefPubMedGoogle Scholar
- 26.Macdonald LE, Karow M, Stevens S, Auerbach W, Poueymirou WT, Yasenchak J, Frendewey D, Valenzuela DM, Giallourakis CC, Alt FW, Yancopoulos GD, Murphy AJ (2013) Precise and in situ genetic humanization of 6 Mb of mouse immunoglobulin genes. Proc Natl Acad Sci U S A 111:5147–5152CrossRefGoogle Scholar
- 30.Stoger E, Schillberg S, Twyman RM, Fischer R, Christou P (2004) Antibody production in transgenic plants. In: Lo BKC (ed) Antibody engineering: protocols and methods, 2nd edn, Methods in molecular biology. Humana Press Inc, Totowa, NJ, pp 301–318Google Scholar