Depixus SAS, 3/5 Impasse Reille, 75014, Paris, France.
Laboratoire de physique de L'École normale supérieure de Paris, CNRS, ENS, Université PSL, Sorbonne Université, Université de Paris, Paris, 75005, France.
Commun Biol. 2021 Jan 29;4(1):128. doi: 10.1038/s42003-021-01648-7.
Accurate decoding of nucleic acid variation is critical to understand the complexity and regulation of genome function. Here we use a single-molecule magnetic tweezer (MT) platform to identify sequence variation and map a range of important epigenetic base modifications with high sensitivity, specificity, and precision in the same single molecules of DNA or RNA. We have also developed a highly specific amplification-free CRISPR-Cas enrichment strategy to isolate genomic regions from native DNA. We demonstrate enrichment of DNA from both E. coli and the FMR1 5'UTR coming from cells derived from a Fragile X carrier. From these kilobase-length enriched molecules we could characterize the differential levels of adenine and cytosine base modifications on E. coli, and the repeat expansion length and methylation status of FMR1. Together these results demonstrate that our platform can detect a variety of genetic, epigenetic, and base modification changes concomitantly within the same single molecules.
准确解码核酸变异对于理解基因组功能的复杂性和调控至关重要。在这里，我们使用单分子磁镊（MT）平台，以高灵敏度、特异性和精度在单个 DNA 或 RNA 分子中识别序列变异，并绘制一系列重要的表观遗传碱基修饰图谱。我们还开发了一种高度特异性的无扩增 CRISPR-Cas 富集策略，用于从天然 DNA 中分离基因组区域。我们展示了来自脆性 X 携带者来源的细胞的大肠杆菌和 FMR1 5'UTR 的 DNA 的富集。从这些千碱基长度的富集分子中，我们可以描述大肠杆菌中腺嘌呤和胞嘧啶碱基修饰的差异水平，以及 FMR1 的重复扩展长度和甲基化状态。这些结果共同表明，我们的平台可以在同一单个分子中同时检测多种遗传、表观遗传和碱基修饰变化。
