Gene Discovery for Bud Dormancy Induction and Release in Oak by 454-Pyrosequencing and Q-PCR Analysis

In temperate regions, the time lag between vegetative bud burst and bud set (dormancy) determines the duration of the growing season of trees. Dormancy allows trees to survive cold injury resulting from exposure to low temperatures. An understanding of the molecular machinery controlling the shift between these two phonological states is of key importance in the context of climatic change. The objective of this study was to identify genes upregulated during endo- and ecodormancy in sessile oak (Quercus petraea) populations located in northwest France. A forcing test on young trees was first carried out to identify the period most likely to correspond to these two stages. We then extracted total RNA from apical buds displaying endo- and ecodormancy. This RNA was used for the generation of cDNA libraries, and used for in-depth transcriptome characterization with 454 FLX pyrosequencing technology, which produced a total of 495,915 reads. Valid reads were then mapped onto oak UniGene data. Digital gene expression analysis was performed on 6,471 contigs with read numbers > 5. The number of sequences displaying significant differences in expression level (read abundance) between endo- and ecodormancy conditions ranged from 48 to 122, depending on algorithms used.  The genes upregulated were enriched in the “response to abiotic stimulus”, “response to stress” and “response to endogenous stimulus” functional categories. We selected 13 genes displaying significant differences between conditions for further analysis, and 11 of these genes, including those for gluthatione-S-transferase (GST) and dehydrin xero2 (XERO2) were validated by quantitative PCR.