Identification of genes involved in ripening of octoploid strawberry (Fragaria x ananassa Duch.) using cDNA-AFLP

Strawberry is a typical non-climacteric fruit, its ripening is not well understood yet, particularly the role of ethylene is unclear. In climacteric fruits, ethylene sets up the process of ripening by activation of ripening-specific gene transcription. Ethylene is present in strawberry too, but its effect is not known yet. Therefore, our aims were to identify genes involved in ripening and ethylene-synthesis, their isolation and characterization in strawberry. For this purpose cDNA-AFLP strategy was used which is able to identify genes showing different expression level. It does not need previous sequence information; it is sensitive, reproducible and quantifiable. This technique is suitable for identifying novel genes and differentiating between homologues. The fruit material (Fragaria x ananassa Duch. cv. 'Elsanta') represent four different ripening stages (green, white, pink and fully ripe) with and without 1-MCP (1-methyl-cyclopropene) treatment. 1-MCP binds specifically to the ethylene receptors blocking the ethylene signal transduction pathway, so it is suitable to find genes indirectly, activated by ethylene. The 270 cDNA fragments amplified by cDNA-AFLP method showed significant difference in expression pattern, according to ripening stage and tissue type. The cDNA fragments were grouped by Cluster and TreeView programs into three categories: genes expressing in the achenes, in the green receptacles and the ripe (red) receptacle. After statistical analysis, cDNA fragments were isolated and re-amplified by PCR. The sequences were compared to data base sequences to find homology. Sequence analysis of cDNA fragments showed, that some of the cDNAs represent genes having key role in ripening, their expression pattern is similar to that of climacteric fruits. Other genes - including genes functioning in the achene - code for proteins: storage, allergic and heat-shock ones.