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Transgenic expression in citrus of single-chain antibody fragments specific to Citrus tristeza virus confers virus resistance

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Abstract

Citrus tristeza virus (CTV) causes one of the most destructive viral diseases of citrus worldwide. Generation of resistant citrus genotypes through genetic engineering could be a good alternative to control CTV. To study whether production of single-chain variable fragment (scFv) antibodies in citrus could interfere and immunomodulate CTV infection, transgenic Mexican lime plants expressing two different scFv constructs, separately and simultaneously, were generated. These constructs derived from the well-referenced monoclonal antibodies 3DF1 and 3CA5, specific against CTV p25 major coat protein, whose mixture is able to detect all CTV isolates characterized so far. ScFv accumulation levels were low and could be readily detected just in four transgenic lines. Twelve homogeneous and vigorous lines were propagated and CTV-challenged by graft inoculation with an aggressive CTV strain. A clear protective effect was observed in most transgenic lines, which showed resistance in up to 40–60% of propagations. Besides, both a delay in symptom appearance and attenuation of symptom intensity were observed in infected transgenic plants compared with control plants. This effect was more evident in lines carrying the 3DF1scFv transgene, being probably related to the biological functions of the epitope recognized by this antibody. This is the first report describing successful protection against a pathogen in woody transgenic plants by ectopic expression of scFv recombinant antibodies.

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Acknowledgments

We thank J.A. Pina and J. Piquer for their technical assistance in growing citrus plants in greenhouses, and P. Serra for his help with western blot analysis. This research was supported by grants Prometeo/2008/121 from the Generalitat Valenciana and AGL2009-08052, RTA2005-00190 and AGL2009-07531 from the Ministerio de Ciencia e Innovación. M. Cervera is recipient of a “Ramón y Cajal” contract. O. Esteban and M. Gil were recipients of PhD fellowships from IVIA.

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  1. Olga Esteban

    Present address: Institut de Bioenginyeria de Catalunya (IBEC), Ed. Hèlix, Baldiri Reixac 15–21, 08028, Barcelona, Spain

Authors and Affiliations

  1. Centro de Protección Vegetal y Biotecnología, Instituto Valenciano de Investigaciones Agrarias (IVIA), Ctra. Moncada-Naquera Km 4.5, 46113, Moncada, Valencia, Spain

    Magdalena Cervera, Olga Esteban, Maite Gil, M. Teresa Gorris, M. Carmen Martínez, Leandro Peña & Mariano Cambra

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  1. Magdalena Cervera
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  2. Olga Esteban
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Correspondence to Mariano Cambra.

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Cervera, M., Esteban, O., Gil, M. et al. Transgenic expression in citrus of single-chain antibody fragments specific to Citrus tristeza virus confers virus resistance. Transgenic Res 19, 1001–1015 (2010). https://doi.org/10.1007/s11248-010-9378-5

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