Alonso Conchita, Pérez Ricardo, Bazaga Pilar, Medrano Mónica, Herrera Carlos M
Estación Biológica de Doñana, CSIC, Avenida Américo Vespucio s/n, Sevilla, 41092, Spain.
Instituto de Investigaciones Químicas, Centro de Investigaciones Científicas Isla de La Cartuja, CSIC-US, Avenida Américo Vespucio s/n, Sevilla, 41092, Spain.
Mol Ecol Resour. 2016 Jan;16(1):80-90. doi: 10.1111/1755-0998.12426. Epub 2015 May 18.
Methylation of DNA cytosines affects whether transposons are silenced and genes are expressed, and is a major epigenetic mechanism whereby plants respond to environmental change. Analyses of methylation-sensitive amplification polymorphism (MS-AFLP or MSAP) have been often used to assess methyl-cytosine changes in response to stress treatments and, more recently, in ecological studies of wild plant populations. MSAP technique does not require a sequenced reference genome and provides many anonymous loci randomly distributed over the genome for which the methylation status can be ascertained. Scoring of MSAP data, however, is not straightforward, and efforts are still required to standardize this step to make use of the potential to distinguish between methylation at different nucleotide contexts. Furthermore, it is not known how accurately MSAP infers genome-wide cytosine methylation levels in plants. Here, we analyse the relationship between MSAP results and the percentage of global cytosine methylation in genomic DNA obtained by HPLC analysis. A screening of literature revealed that methylation of cytosines at cleavage sites assayed by MSAP was greater than genome-wide estimates obtained by HPLC, and percentages of methylation at different nucleotide contexts varied within and across species. Concurrent HPLC and MSAP analyses of DNA from 200 individuals of the perennial herb Helleborus foetidus confirmed that methyl-cytosine was more frequent in CCGG contexts than in the genome as a whole. In this species, global methylation was unrelated to methylation at the inner CG site. We suggest that global HPLC and context-specific MSAP methylation estimates provide complementary information whose combination can improve our current understanding of methylation-based epigenetic processes in nonmodel plants.
DNA胞嘧啶甲基化影响转座子是否沉默以及基因是否表达，是植物响应环境变化的主要表观遗传机制。甲基化敏感扩增多态性分析（MS-AFLP或MSAP）常被用于评估应激处理后甲基胞嘧啶的变化，最近也用于野生植物种群的生态学研究。MSAP技术不需要已测序的参考基因组，可提供许多随机分布在基因组上的匿名位点，通过这些位点可以确定甲基化状态。然而，MSAP数据的评分并不简单，仍需努力规范这一步骤，以利用区分不同核苷酸背景下甲基化的潜力。此外，尚不清楚MSAP在推断植物全基因组胞嘧啶甲基化水平方面的准确性如何。在此，我们分析了MSAP结果与通过高效液相色谱（HPLC）分析获得的基因组DNA中全局胞嘧啶甲基化百分比之间的关系。文献筛选表明，MSAP检测的切割位点处胞嘧啶的甲基化程度高于HPLC获得的全基因组估计值，不同核苷酸背景下的甲基化百分比在物种内和物种间存在差异。对多年生草本植物臭嚏根草200个个体的DNA同时进行HPLC和MSAP分析证实，甲基胞嘧啶在CCGG背景中比在整个基因组中更常见。在该物种中，全局甲基化与内部CG位点的甲基化无关。我们认为，全局HPLC和特定背景的MSAP甲基化估计提供了互补信息，将二者结合可以增进我们目前对非模式植物中基于甲基化的表观遗传过程的理解。
