Analysis of gene-targeted DNA methylation patterns through the annual cycle in clonal epitypes of Norway spruce

Main goal of the study is to distinguish stable DNA methylation differences from dynamic changes during the annual cycle in different epitypes. We examined whether the extant epigenetic marks created during embryogenesis under EpI-temperature conditions are maintained in the resulting epitype trees. We used unique well-defined clonal epitypes with a stable epigenetic memory of the temperature sum experienced during embryogenesis impacting bud phenology. Due to the massive size of the Norway spruce genome (Nystedt et al., 2013), we used a targeted approach to address the DNA methylation status at a single-base resolution in specific target gene regions of selected genes involved in the epigenetic machinery and adaptive traits formation. From these we have characterized DNA methylation patterns in apical buds for 10 time points throughout the year to observe the differences between epitypes and its dynamics through the annual cycle of development. 60 pooled captured bisulfite-converted DNA libraries were sequenced at the BGI Genomics (Hong Kong, China) using DNBSEQ (G-400) sequencing platform with 150PE with 40% in lane control sequencing.

本研究的核心目标是区分不同表观型（epitype）在年度周期中稳定存在的DNA甲基化差异与动态变化。我们旨在探究胚胎发生阶段于EpI-温度条件下形成的现存表观遗传标记，是否会在由此产生的表观型树木中得以保留。我们选用了具有稳定表观遗传记忆的独特且经明确定义的克隆表观型，该表观型的表观遗传记忆可反映胚胎发生阶段所经历的积温对芽物候的影响。鉴于挪威云杉基因组规模庞大（Nystedt等，2013），我们采用靶向测序策略，在参与表观遗传调控机制与适应性性状形成的选定基因的特定靶区域内，以单碱基分辨率解析DNA甲基化状态。基于上述实验设计，我们对全年10个时间点的顶芽进行了DNA甲基化模式表征，以观察不同表观型之间的差异及其在年度发育周期中的动态变化。本研究共构建60个混合捕获亚硫酸氢盐转化DNA（bisulfite-converted DNA）文库，并委托中国香港的BGI Genomics使用DNBSEQ（G-400）测序平台开展150PE双端测序，且每个泳道设置40%的对照测序样本。