通过纳米限域电化学发光平台对随机 DNA 序列上的 5-甲基胞嘧啶和 5-羟甲基胞嘧啶进行鉴定和定量,Analytical Chemistry

当前位置： X-MOL 学术 › Anal. Chem. › 论文详情

Our official English website, www.x-mol.net, welcomes your feedback! (Note: you will need to create a separate account there.)

通过纳米限域电化学发光平台对随机 DNA 序列上的 5-甲基胞嘧啶和 5-羟甲基胞嘧啶进行鉴定和定量
Analytical Chemistry ( IF 6.7 ) Pub Date : 2023-06-13 , DOI: 10.1021/acs.analchem.3c01252
Mao-Hua Gao ₁ , Mei-Chen Pan ₁ , Pu Zhang ₁ , Wen-Bin Liang ₁ , Xia Zhong ₁ , Ying Zhuo ₁

Affiliation

5-甲基胞嘧啶（5mC）和5-羟甲基胞嘧啶（5hmC）是哺乳动物基因组中最丰富的两种表观遗传标记，并且已证明这些双重表观遗传标记比单个标记能够更准确地预测癌症的复发和生存。然而，由于5mC和5hmC的结构相似且表达量低，区分和定量这两种甲基化修饰具有挑战性。在此，我们使用 10-11 易位家族双加氧酶 (TET)通过以下方式将 5mC 转化为 5hmC：基于纳米限域电化学发光（ECL）平台结合重组酶聚合酶扩增（RPA）辅助CRISPR/Cas13a系统的扩增策略，实现了两种标记的识别。受益于TET介导的转换策略，开发了高度一致的标记途径来识别随机序列上的双表观遗传标记，有效减少了系统误差。通过制备碳化聚合物点嵌入SiO ₂纳米网络(CPDs@SiO ₂），由于纳米约束增强的 ECL 效应，与散射发射器相比，它表现出更高的 ECL 效率和更稳定的 ECL 性能。所提出的生物分析策略可分别用于鉴定和定量 100 aM 至 100 pM 范围内的 5mC 和 5hmC，这为早期诊断与异常甲基化相关的疾病提供了一种有前途的工具。

"点击查看英文标题和摘要"

Identification and Quantification of 5-Methylcytosine and 5-Hydroxymethylcytosine on Random DNA Sequences by a Nanoconfined Electrochemiluminescence Platform

5-Methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) are two of the most abundant epigenetic marks in mammalian genomes, and it has been proven that these dual epigenetic marks give a more accurate prediction of recurrence and survival in cancer than the individual mark. However, due to the similar structure and low expression of 5mC and 5hmC, it is challenging to distinguish and quantify the two methylation modifications. Herein, we employed the ten-eleven translocation family dioxygenases (TET) to convert 5mC to 5hmC via a specific labeling process, which realized the identification of the two marks based on a nanoconfined electrochemiluminescence (ECL) platform combined with the amplification strategy of a recombinase polymerase amplification (RPA)-assisted CRISPR/Cas13a system. Benefiting from the TET-mediated conversion strategy, a highly consistent labeling pathway was developed for identifying dual epigenetic marks on random sequence, which reduced the system error effectively. The ECL platform was established via preparing a carbonized polymer dot embedded SiO₂ nanonetwork (CPDs@SiO₂), which exhibited higher ECL efficiencies and more stable ECL performance compared to those of the scattered emitters due to the nanoconfinement-enhanced ECL effect. The proposed bioanalysis strategy could be employed for the identification and quantification of 5mC and 5hmC in the range from 100 aM to 100 pM, respectively, which provides a promising tool for early diagnosis of diseases associated with abnormal methylation.

更新日期：2023-06-13

点击分享查看原文

点击收藏

阅读更多本刊新发论文本刊介绍/投稿指南