New England Biolabs, Incorporated, Ipswich, Massachusetts 01938, USA.
SLC Management, Wellesley Hills, Massachusetts 02481, USA.
Genome Res. 2022 Nov-Dec;32(11-12):2079-2091. doi: 10.1101/gr.277080.122. Epub 2022 Nov 4.
Covalent modifications of genomic DNA are crucial for most organisms to survive. Amplicon-based high-throughput sequencing technologies erase all DNA modifications to retain only sequence information for the four canonical nucleobases, necessitating specialized technologies for ascertaining epigenetic information. To also capture base modification information, we developed Methyl-SNP-seq, a technology that takes advantage of the complementarity of the double helix to extract the methylation and original sequence information from a single DNA molecule. More specifically, Methyl-SNP-seq uses bisulfite conversion of one of the strands to identify cytosine methylation while retaining the original four-bases sequence information on the other strand. As both strands are locked together to link the dual readouts on a single paired-end read, Methyl-SNP-seq allows detecting the methylation status of any DNA even without a reference genome. Because one of the strands retains the original four nucleotide composition, Methyl-SNP-seq can also be used in conjunction with standard sequence-specific probes for targeted enrichment and amplification. We show the usefulness of this technology in a broad spectrum of applications ranging from allele-specific methylation analysis in humans to identification of methyltransferase specificity in complex bacterial communities.
共价修饰的基因组 DNA 是至关重要的，大多数生物体生存。基于扩增子的高通量测序技术擦除所有的 DNA 修饰只保留序列信息的四个典型核碱基，需要专门的技术来确定表观遗传信息。为了捕获碱基修饰信息，我们开发了甲基 SNP-seq，一种利用双链互补性从单个 DNA 分子中提取甲基化和原始序列信息的技术。更具体地说，甲基 SNP-seq 使用亚硫酸氢盐转化的一条链来识别胞嘧啶甲基化，同时保留另一条链上的原始四碱基序列信息。由于两条链被锁定在一起，将单个配对末端读取的双读取结果联系起来，甲基 SNP-seq 允许检测任何 DNA 的甲基化状态，即使没有参考基因组。由于其中一条链保留了原始的四个核苷酸组成，甲基 SNP-seq 也可以与标准的序列特异性探针一起用于靶向富集和扩增。我们展示了这项技术在广泛的应用中的有用性，从人类的等位基因特异性甲基化分析到复杂细菌群落中甲基转移酶特异性的鉴定。
