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P-109<br />

Characterization of an ankyrin-like protein up-stream region from Vitis vinifera is highly<br />

induced by Botrytis cinerea infection<br />

M.A. Miccono 1 , M. Ortega 2 , C. Montes 1 , P. Barba* 1 , J. Rubio 1 , H. Peña-Cortés 3 , H. Prieto 1<br />

1 Biotechnology Laboratory, INIA-La Platina Station, La Platina, Chile; 2 Doctoral Program in<br />

Biotechnology, University of Santiago, Santiago, Chile; 3 D. Alkalay L. Biotechnology Center,<br />

Federico Santa María University, Valparaíso, Chile<br />

*Corresponding author: pbarba@gmail.com<br />

Grey mold, caused by Botrytis cinerea, is a main grapevine disease in Chile. B. cinerea is a<br />

necrotrophic filamentous fungus of a complex agricultural management because of a broad host<br />

spectrum <strong>and</strong> to the different sources of inoculation. In the “Genome I: Botrytis-Grape<br />

Interaction” project, a macroarray formed by 4803 ESTs was designed. Gene expression studies<br />

using this macroarray allowed the comparison between mRNAs from infected <strong>and</strong> non-infected<br />

grapevine field plants from two contrasting cultivars: Thompson Seedless <strong>and</strong> Carménère. By<br />

this analysis, we established the involvement of a putative ankyrin-like protein highly induced in<br />

response to the pathogen infection. Although is not expected to find natural resistance sources to<br />

Botrytis in breeding grapevine studies using susceptible cultivars, the knowledge of highly<br />

responsible sequences to fungal challenge could be of significant relevance from a<br />

biotechnological approach. In this way, we propose that the promoter region of this gene may<br />

contain important responsive element which may provide information about the specificity of the<br />

plant response after fungal attack. A first step in this work was to verify <strong>and</strong> compare the<br />

observed induction of the grapevine ankyrin-like gene under fungal attack by specific real time<br />

PCR analysis of infected Carménère greenhouse plants. Afterwards, in silico analysis was carried<br />

out leading to the definition of an 890 base pair up-stream zone, which was experimentally<br />

characterized by use of GFP fusion constructs in tobacco transformation experiments. Wounding<br />

<strong>and</strong> Botrytis challenges were applied on stable transgenic lines, depicting differentially induced<br />

levels of gfp transcripts depending on the elicitor. Comparison of these heterologous inductions<br />

is under development by generation of the corresponding ´Thompson Seedless´ transgenic lines.<br />

190


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