Liau Yusmiati, Whibley Annabel, Hill Amy M, Vanga Bhanupratap R, Pither-Joyce Meeghan, Hilario Elena, Bailey Sarah, Thomson Susan J, Lizamore Darrell
Grapevine Improvement, Bragato Research Institute, Lincoln, New Zealand.
School of Biological Sciences, University of Auckland, Auckland, New Zealand.
BMC Genomics. 2024 Dec 23;25(1):1235. doi: 10.1186/s12864-024-11145-w.
Nanopore sequencing enables detection of DNA methylation at the same time as identification of canonical sequence. A recent study validated low-pass nanopore sequencing to accurately estimate global methylation levels in vertebrates with sequencing coverage as low as 0.01x. We investigated the applicability of this approach to plants by testing three plant species and analysed the effect of technical and biological parameters on estimate precision and accuracy. Our results indicate that higher coverage (0.1x) is required to achieve accuracy in assessing plant global methylation comparable to that in vertebrates. Shorter read length and a closer sequence match between sample and reference genome improved measurement accuracy. Application of this method in Vitis vinifera showed consistent global methylation levels across different leaf sizes, and different sample preservation and DNA extraction methods, whereas different varieties and tissue types did exhibit methylation differences. Similarly, distinct methylation patterns were observed in different genomic features. Our findings suggest the suitability of this method as a low-cost screening tool for validation of experimental parameters, developmental time courses, and to assess methylation status for different modification types and sequence contexts at the level of whole genome or for abundant genomic features such as transposable elements.
纳米孔测序能够在识别标准序列的同时检测DNA甲基化。最近的一项研究验证了低通量纳米孔测序可在测序覆盖度低至0.01x的情况下准确估计脊椎动物的整体甲基化水平。我们通过测试三种植物物种研究了该方法对植物的适用性，并分析了技术和生物学参数对估计精度和准确性的影响。我们的结果表明，要在评估植物整体甲基化方面达到与脊椎动物相当的准确性，需要更高的覆盖度（0.1x）。较短的读长以及样本与参考基因组之间更紧密的序列匹配提高了测量准确性。该方法在葡萄中的应用表明，不同叶片大小、不同样本保存和DNA提取方法下的整体甲基化水平是一致的，而不同品种和组织类型确实存在甲基化差异。同样，在不同的基因组特征中观察到了明显的甲基化模式。我们的研究结果表明，该方法适合作为一种低成本的筛选工具，用于验证实验参数、发育时间进程，以及在全基因组水平或对于转座元件等丰富的基因组特征评估不同修饰类型和序列背景下的甲基化状态。
