meHOLMES: A CRISPR-cas12a-based method for rapid detection of DNA methylation in a sequence-independent manner

Aberrant DNA methylation is closely associated with various diseases, particularly cancer, and its precise detection plays an essential role in disease diagnosis and monitoring. In this study, we present a novel DNA methylation detection method (namely meHOLMES), which integrates both the TET2/APOBE...

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Bibliographic Details
Published in:Heliyon 2024-01, Vol.10 (2), p.e24574-e24574, Article e24574
Main Authors: Zhuang, Songkuan, Hu, Tianshuai, Zhou, Xike, Zhou, Hongzhong, He, Shiping, Li, Jie, Qiu, Long, Zhang, Yuehui, Xu, Yong, Pei, Hao, Gu, Dayong, Wang, Jin
Format: Article
Language:English
Subjects:
APOBEC
cost effectiveness
CRISPR-Cas12a
cytosine
deamination
disease diagnosis
DNA methylation
fluorescence
HOLMES
immunoaffinity chromatography
rapid methods
TET2
thymine
uracil
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Summary:Aberrant DNA methylation is closely associated with various diseases, particularly cancer, and its precise detection plays an essential role in disease diagnosis and monitoring. In this study, we present a novel DNA methylation detection method (namely meHOLMES), which integrates both the TET2/APOBEC-mediated cytosine deamination step and the CRISPR-Cas12a-based signal readout step. TET2/APOBEC efficiently converts unmethylated cytosine to uracil, which is subsequently changed to thymine after PCR amplification. Utilizing a rationally designed crRNA, Cas12a specifically identifies unconverted methylated cytosines and generates detectable signals using either fluorescent reporters or lateral flow test strips. meHOLMES quantitatively detects methylated CpG sites with or without Protospacer Adjacent Motif (PAM) sequences in both artificial and real biological samples. In addition, meHOLMES can complete the whole detection process within 6 h, which is much faster than traditional bisulfite-based sample pre-treatment method. Above all, meHOLMES provides a simpler, faster, more accurate, and cost-effective approach for quantitation of DNA methylation levels in a sequence-independent manner. •MeHOLMES conveniently detects CpG methylation with either fluorescence or LFA readout.•MeHOLMES completes the whole CpG methylation detection process within 6 h.•MeHOLMES detects CpG methylation levels in a sequence-independent manner.•MeHOLMES quantitatively detects CpG methylation levels in real biological samples.•MeHOLMES empolys milder and simpler sample pretreatment through using TET2/APOBEC.
ISSN:2405-8440
2405-8440
DOI:10.1016/j.heliyon.2024.e24574