LongSAGE transcriptome comparison on Botrytis cinerea conidia germination with and without resveratrol treatment

Two long serial analysis of gene expression (LongSAGE) libraries of germinating conidia of Botrytis cinerea were generated with and without resveratrol treatments for unraveling the antifungal mechanisms of this phytoalexin by global transcriptome analysis – 82,786 total tags and a minimum of 15,665 unitags were obtained respectively. By tag-to-gene mapping, about 55% of the total unitags were matched to known genes. There were 109 unitags that showed significant differences in the two LongSAGE libraries, corresponding to 43 up- and 44 down-regulated genes, and 22 unmatched unitags as new gene candidates. These results showed that resveratrol inhibited expression of genes related to the glycolysis-Krebs cycle and mitochondrial electron transport chains at multiple sites in conidia germination, with induction of modified Entner–Doudoroff pathway as metabolic compensation. The chaperone-mediated protein folding and ubiquitin proteolysis systems were inhibited, while detoxification of resveratrol via ABC transporters was activated in this fungal pathogen. Keywords: Botrytis cinerea; LongSAGE; metabolic pathway; resveratrol; transcriptome Botrytis cinerea Pers. Ex was identified and isolated from stage III fruit of grapevines (Vitis vinifera cv. Merlot) in Huailai, Beijing. The most highly pathogenic strain M4 was chosen for these experiments. PSA (300g potato, 20g sugar, 20g agar per liter) was used for Botrytis culture at 20-23°C (Adrian et al. 1997). Conidia were aseptically harvested from 14 d cultures by washing with PSM (37.5g potato, 20g sugar per liter). Conidia treatments and germination: 5 x 105 cfu/ml conidia in PSM was added with resveratrol (for BCB library treatment) to a final concentration of 140 µg/ml tran-resveratrol (Sigma, >99% purity), with no addition of resveratrol as a control (for BCA library treatment) (Adrian et al. 1997). After 12-14 h incubation in the dark at 21-22°C with shaking in shallow Petri dishes, 5 x 106 conidia from each treatment were collected and washed with H2O by centrifugation at 1,600g for 20 min at 21°C. Conidia were then frozen and stored at -80°C for RNA isolation.