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Whole-genome long-read TAPS deciphers DNA methylation patterns at base resolution using PacBio SMRT sequencing technology
Nucleic Acids Research ( IF 16.6 ) Pub Date : 2022-07-18 , DOI: 10.1093/nar/gkac612
Jinfeng Chen 1, 2 , Jingfei Cheng 1, 2 , Xiufei Chen 1, 2 , Masato Inoue 1, 2 , Yibin Liu 1, 2 , Chun-Xiao Song 1, 2
Affiliation  

Long-read sequencing provides valuable information on difficult-to-map genomic regions, which can complement short-read sequencing to improve genome assembly, yet limited methods are available to accurately detect DNA methylation over long distances at a whole-genome scale. By combining our recently developed TET-assisted pyridine borane sequencing (TAPS) method, which enables direct detection of 5-methylcytosine and 5-hydroxymethylcytosine, with PacBio single-molecule real-time sequencing, we present here whole-genome long-read TAPS (wglrTAPS). To evaluate the performance of wglrTAPS, we applied it to mouse embryonic stem cells as a proof of concept, and an N50 read length of 3.5 kb is achieved. By sequencing wglrTAPS to 8.2× depth, we discovered a significant proportion of CpG sites that were not covered in previous 27.5× short-read TAPS. Our results demonstrate that wglrTAPS facilitates methylation profiling on problematic genomic regions with repetitive elements or structural variations, and also in an allelic manner, all of which are extremely difficult for short-read sequencing methods to resolve. This method therefore enhances applications of third-generation sequencing technologies for DNA epigenetics.

中文翻译:


全基因组长读长 TAPS 使用 PacBio SMRT 测序技术以碱基分辨率破译 DNA 甲基化模式



长读长测序提供了有关难以绘制基因组区域的宝贵信息,可以补充短读长测序以改善基因组组装,但可用于在全基因组规模上长距离准确检测 DNA 甲基化的方法有限。通过将我们最近开发的 TET 辅助吡啶硼烷测序 (TAPS) 方法(可直接检测 5-甲基胞嘧啶和 5-羟甲基胞嘧啶)与 PacBio 单分子实时测序相结合,我们在此推出全基因组长读长 TAPS( wglrTAPS)。为了评估 wglrTAPS 的性能,我们将其应用于小鼠胚胎干细胞作为概念证明,并实现了 3.5 kb 的 N50 读长。通过对 wglrTAPS 进行 8.2 倍深度的测序,我们发现了之前 27.5 倍短读长 TAPS 中未覆盖的很大一部分 CpG 位点。我们的结果表明,wglrTAPS 有助于对具有重复元件或结构变异的有问题的基因组区域进行甲基化分析,并且还以等位基因的方式进行甲基化分析,所有这些对于短读长测序方法来说都极难解决。因此,该方法增强了第三代测序技术在 DNA 表观遗传学中的应用。
更新日期:2022-07-18
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