通过组合索引和酶促DNA甲基化转化进行单细胞DNA甲基化测序。

Single-cell DNA methylation sequencing by combinatorial indexing and enzymatic DNA methylation conversion.

作者信息

Chatterton Zac, Lamichhane Praves, Ahmadi Rastegar Diba, Fitzpatrick Lauren, Lebhar Hélène, Marquis Christopher, Halliday Glenda, Kwok John B

机构信息

Brain and Mind Centre, The University of Sydney, Camperdown, Australia.

School of Medical Science, The University of Sydney, Camperdown, Australia.

出版信息

Cell Biosci. 2023 Jan 4;13(1):2. doi: 10.1186/s13578-022-00938-9.

DOI:10.1186/s13578-022-00938-9

PMID:36600255

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC9811750/

Abstract

BACKGROUND

DNA methylation is a critical molecular mark involved in cellular differentiation and cell-specific processes. Single-cell whole genome DNA methylation profiling methods hold great potential to resolve the DNA methylation profiles of individual cell-types. Here we present a method that couples single-cell combinatorial indexing (sci) with enzymatic conversion (sciEM) of unmethylated cytosines.

RESULTS

The sciEM method facilitates DNA methylation profiling of single-cells that is highly correlated with single-cell bisulfite-based workflows (r > 0.99) whilst improving sequencing alignment rates, reducing adapter contamination and over-estimation of DNA methylation levels (CpG and non-CpG). As proof-of-concept we perform sciEM analysis of the temporal lobe, motor cortex, hippocampus and cerebellum of the human brain to resolve single-cell DNA methylation of all major cell-types.

CONCLUSION

To our knowledge sciEM represents the first non-bisulfite single-cell DNA methylation sequencing approach with single-base resolution.

摘要

背景

DNA甲基化是一种参与细胞分化和细胞特异性过程的关键分子标记。单细胞全基因组DNA甲基化分析方法在解析单个细胞类型的DNA甲基化图谱方面具有巨大潜力。在此，我们介绍一种将单细胞组合索引（sci）与未甲基化胞嘧啶的酶促转化（sciEM）相结合的方法。

结果

sciEM方法有助于对单细胞进行DNA甲基化分析，其结果与基于亚硫酸氢盐的单细胞工作流程高度相关（r > 0.99），同时提高了测序比对率，减少了接头污染，并降低了DNA甲基化水平（CpG和非CpG）的高估。作为概念验证，我们对人类大脑的颞叶、运动皮层、海马体和小脑进行了sciEM分析，以解析所有主要细胞类型的单细胞DNA甲基化情况。