NaCl胁迫对黄瓜种子萌发的影响及DNA甲基化的MSAP分析

Effects of NaCl Stress on Seed Germination and DNA Methylation Status Detected by MSAP Analysis in Cucumber

【目的】研究黄瓜种子萌发过程中DNA甲基化动态变化情况以及外源施加NaCl对种子萌发及种子DNA甲基化水平的影响,探索DNA甲基化在黄瓜种子萌发过程及NaCl胁迫反应中的作用。【方法】运用甲基化敏感扩增多态性(methylation se nsitive amplified polymorphism,MSAP)技术,分析0、150、200 mmol.L-1 NaCl处理下,黄瓜种子萌发过程(0、1、2、4、6、8 d)的DNA甲基化水平及动态变化情况。【结果】MSAP结果显示,黄瓜种子CCGG位点的甲基化水平约15.25%,以双链甲基化方式为主。种子中DNA甲基化水平在萌发1—2 d略有升高,而在整个萌发过程中呈下降趋势;NaCl处理加剧了甲基化的变化幅度,并使萌发末期的甲基化水平更低。萌发过程中甲基化升高和降低同时发生,分别在不同萌发时期占主导,不同变化类型中CG/CHG(H=A,T,C)位点胞嘧啶甲基化同时变化的类型占主导。甲基化变化同时发生在编码序列和非编码序列中。【结论】黄瓜种子萌发过程的DNA甲基化表现出复杂性,甲基化和去甲基化进程同时进行,并具有时空特异性。NaCl处理降低了种子基因组甲基化水平的稳定性,对种子萌发有抑制作用。

[ Objective ] The objectives of this study were to analyze the cytosine methylation dynamics during cucumber seed germination and to assess the effect of NaC1 treatment on seed germination and DNA methylation status. Moreover, the role of DNA methylation on seed germination and salt stress adaptation was studied at the same time. [ Method] Methylation sensitive amplified polymorphism （MSAP） method was used to analyze the level and variation patterns ofDNA methylation at different times （0, 1, 2, 4, 6, 8 d） during cucumber seed germination in different NaCl treatments （0, 150, 200 mmol·L-1）.[Result]MSAP results showed that the methylation level in cucumber dry seeds was 15.25%, mainly in full methylation pattern. The level of global DNA methylation decreased during the whole germination process, with a slight increase at the early stages （1-2 d）. NaCl treatment aggravated these changes, and made the final level of DNA methylation lower. Hypermethylation and hypomethylation happened simultaneously, and predominated at different germination stages. Among different change types, CG/CHG （H=A, T, C） simultaneous changes were the dominating types. DNA methylation changes were detected in both coding region and non-coding region on the cucumber genome. [ Conclusion ] DNA methylation changes during seed germination is a complex process. Methylation and demethylation happened at the same time in a temporal-spatial specific way. NaC1 treatment reduced the cucumber genome stability and inhibited the seed germination.