DNA modifications in bacteria present diverse types and distributions, playing crucial functional roles. Current methods for detecting bacterial DNA modifications via nanopore sequencing typically involve comparing raw current signals to a methylation-free control. In this study, we found that bacterial DNA modification induces errors in nanopore reads. And these errors are found only in one strand but not the other, showing a strand-specific bias. Leveraging this discovery, we developed Hammerhead, a pioneering pipeline designed for de novo methylation discovery that circumvents the necessity of raw signal inference and a methylation-free control. The majority (14 out of 16) of the identified motifs can be validated by raw signal comparison methods or by identifying corresponding methyltransferases in bacteria. Additionally, we included a novel polishing strategy employing duplex reads to correct modification-induced errors in bacterial genome assemblies, achieving a reduction of over 85% in such errors. In summary, Hammerhead enables users to effectively locate bacterial DNA methylation sites from nanopore FASTQ/FASTA reads, thus holds promise as a routine pipeline for a wide range of nanopore sequencing applications, such as genome assembly, metagenomic binning, decontaminating eukaryotic genome assemblies, and functional analysis for DNA modifications.

中文翻译：

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纳米孔链特异性错配能够从头检测细菌 DNA 修饰


细菌中的 DNA 修饰呈现不同的类型和分布，发挥着至关重要的功能作用。目前通过纳米孔测序检测细菌 DNA 修饰的方法通常涉及将原始电流信号与无甲基化对照进行比较。在这项研究中，我们发现细菌 DNA 修饰会诱导纳米孔读数的错误。这些错误仅在一条链中发现，而在另一条链中未发现，显示出特定于链的偏差。利用这一发现，我们开发了 Hammerhead，这是一种专为从头甲基化发现而设计的开创性管道，可避免原始信号推断和无甲基化对照的必要性。大多数 （16 个中的 14 个） 已鉴定的基序可以通过原始信号比较方法或通过鉴定细菌中相应的甲基转移酶来验证。此外，我们还采用了一种新的精纯策略，采用双链读长来纠正细菌基因组组装中修饰诱导的错误，从而将此类错误减少了 85% 以上。总之，Hammerhead 使用户能够从纳米孔 FASTQ/FASTA 读数中有效地定位细菌 DNA 甲基化位点，因此有望作为各种纳米孔测序应用的常规管道，例如基因组组装、宏基因组分箱、去污真核基因组组装和 DNA 修饰的功能分析。