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为了解黑麦草(Lolium perenne)种子在正常条件与NaCl胁迫下萌发过程中DNA甲基化动态变化以及重要盐胁迫相关基因的表达。运用甲基化敏感扩增多态性技术分析对照和150 mmol·L~(-1)NaCl处理下黑麦草种子萌发过程(0,1,2,4,7 d)的DNA甲基化水平及动态变化,并通过实时荧光定量PCR(QRT-PCR)检测8个重要的盐胁迫相关基因的表达情况。结果表明:黑麦草种子萌发过程中甲基化水平呈下降趋势,以双链甲基化方式为主,甲基化变化同时发生在编码序列和非编码序列中。NaCl处理下DNA甲基化程度高于CK,而去甲基化程度低于CK,最终导致NaCl处理下基因组净的甲基化位点数目增加。黑麦草种子耐盐萌发过程中代谢增强,8个盐胁迫相关基因表达量呈上升趋势,而NaCl胁迫延缓了种子萌发进程,在大多数时间点的基因表达低于对照。
In order to understand the dynamic changes of DNA methylation and the expression of important salt stress related genes during the germination process of Lolium perenne seed under normal conditions and NaCl stress. The methylation-sensitive amplification polymorphism (PCR-RFLP) technique was used to analyze the DNA methylation levels of the seeds of ryegrass (0, 1, 2, 4 and 7 d) in the control and 150 mmol·L -1 NaCl treatments Dynamic changes, and detected by real-time quantitative PCR (QRT-PCR) eight important salt stress-related gene expression. The results showed that the methylation level of ryegrass decreased with the double-stranded methylation pattern, and the methylation changed both in coding and non-coding sequences. Under NaCl stress, DNA methylation was higher than that of CK and the degree of demethylation was lower than that of CK, leading to an increase in the number of net methylation sites in the genome under NaCl treatment. The metabolism of ryegrass seeds increased during salt-tolerant germination, and the expression level of eight salt-stress related genes increased while NaCl stress delayed the seed germination process, and the gene expression at most of the time points was lower than that of the control.