Forestry and Forest Products Research Institute, Department of Forest Genetics, Tree Genetics Laboratory, 1 Matsunosato, Tsukuba, Ibaraki 305-8687 Japan.
BMC Genomics. 2013 Apr 10;14:236. doi: 10.1186/1471-2164-14-236.
In temperate regions, the time lag between vegetative bud burst and bud set determines the duration of the growing season of trees (i.e. the duration of wood biomass production). Dormancy, the period during which the plant is not growing, allows trees to avoid cold injury resulting from exposure to low temperatures. An understanding of the molecular machinery controlling the shift between these two phenological 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, the two main stages of bud dormancy. Sessile oak is a widely distributed European white oak species. A forcing test on young trees was first carried out to identify the period most likely to correspond to these two stages. Total RNA was then extracted from apical buds displaying endo- and ecodormancy. This RNA was used for the generation of cDNA libraries, and in-depth transcriptome characterization was performed with 454 FLX pyrosequencing technology.
Pyrosequencing produced a total of 495,915 reads. The data were cleaned, duplicated reads removed, and sequences were mapped onto the oak UniGene data. Digital gene expression analysis was performed, with both R statistics and the R-Bioconductor packages (edgeR and DESeq), on 6,471 contigs with read numbers ≥ 5 within any contigs. The number of sequences displaying significant differences in expression level (read abundance) between endo- and ecodormancy conditions ranged from 75 to 161, depending on the algorithm used. 13 genes displaying significant differences between conditions were selected for further analysis, and 11 of these genes, including those for glutathione-S-transferase (GST) and dehydrin xero2 (XERO2) were validated by quantitative PCR.
The identification and functional annotation of differentially expressed genes involved in the "response to abscisic acid", "response to cold stress" and "response to oxidative stress" categories constitutes a major step towards characterization of the molecular network underlying vegetative bud dormancy, an important life history trait of long-lived organisms.
在温带地区，营养芽爆发和芽形成之间的时间滞后决定了树木的生长季节（即木材生物质生产的持续时间）。休眠是植物不生长的时期，它使树木能够避免因暴露在低温下而受到冷害。了解控制这两个物候状态之间转变的分子机制对于气候变化的背景下至关重要。本研究的目的是鉴定在芽休眠的两个主要阶段，即内休眠和外休眠期间上调的基因。无梗花栎是一种广泛分布的欧洲白栎物种。首先对幼树进行强制试验，以确定最有可能对应于这两个阶段的时期。然后从显示内休眠和外休眠的顶芽中提取总 RNA。使用该 RNA 生成 cDNA 文库，并使用 454 FLX 焦磷酸测序技术进行深度转录组特征分析。
焦磷酸测序共产生了 495915 条读长。对数据进行了清理、去除重复读长，并将序列映射到栎属 UniGene 数据上。使用 R 统计和 R-Bioconductor 包（edgeR 和 DESeq）对 6471 个在任何连续体中读长数≥5 的连续体进行数字基因表达分析。在休眠和休眠条件下，表达水平（读长丰度）差异显著的序列数在 75 到 161 之间，具体取决于所使用的算法。选择了 13 个在条件之间显示显著差异的基因进行进一步分析，其中包括谷胱甘肽-S-转移酶（GST）和脱水素 xero2（XERO2）的 11 个基因，通过定量 PCR 进行了验证。
鉴定和功能注释参与“响应脱落酸”、“响应冷胁迫”和“响应氧化胁迫”类别的差异表达基因，是对营养芽休眠的分子网络进行特征描述的重要步骤，这是长寿生物的一个重要生活史特征。
