Department of Plant and Microbial Biology & Zurich-Basel Plant Science Center, University of Zurich, Zurich, Switzerland.
Methods Mol Biol. 2025;2873:3-17. doi: 10.1007/978-1-0716-4228-3_1.
Whole-genome bisulfite sequencing (WGBS) is the most widely used method to study DNA methylation profiles across the genome. Since the bisulfite reaction causes DNA degradation, a new approach called post-bisulfite adapter tagging (PBAT) was developed to overcome this problem by adding adapters after bisulfite treatment. In mammals, the PBAT method is used for single-cell bisulfite sequencing (scBS-seq), which enables DNA methylation analysis using a very small amount of DNA from only a few cells, including single-cell input. This protocol involves bisulfite conversion, followed by preamplification and tagging with random hexamer primers prior to Illumina library preparation. Since many procedures are completed in one single test tube, the loss of DNA can be minimized, enabling highly sensitive experiments to study DNA methylation profiles from a very small amount of input material.
全基因组亚硫酸氢盐测序（WGBS）是研究全基因组 DNA 甲基化图谱最广泛使用的方法。由于亚硫酸氢盐反应会导致 DNA 降解，因此开发了一种新的方法，称为亚硫酸氢盐处理后接头标记（PBAT），通过在亚硫酸氢盐处理后添加接头来克服这个问题。在哺乳动物中，PBAT 方法用于单细胞亚硫酸氢盐测序（scBS-seq），它可以使用非常少量的 DNA （仅来自几个细胞），包括单细胞输入，来进行 DNA 甲基化分析。该方案包括亚硫酸氢盐转化，然后在 Illumina 文库制备之前进行预扩增和随机六聚体引物标记。由于许多步骤都在一个单独的试管中完成，因此可以最大限度地减少 DNA 的损失，从而能够进行高灵敏度的实验，从非常少量的输入材料中研究 DNA 甲基化图谱。
