Universal Enrichment and Sequencing of DNA Modifications Using a Click-Coupled IgG-Protein G System
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ABSTRACT:
PROVIDER: PRJNA1306169 | ENA |
REPOSITORIES: ENA
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Project description:Efficient nucleic acid enrichment is pivotal for deciphering epigenetic modifications and disease biomarkers, yet current methods are constrained by insufficient specificity, poor versatility, and high costs. We developed a universal strategy named ‘Click-IP-Seq’ by leveraging the high-affinity binding between Protein G and Fc region of DBCO-modified IgG. This enabled the directional conjugation of DBCO-IgG with azide-modified nucleic acids via copper-free strain-promoted azide-alkyne cycloaddition (SPAAC) click chemistry, achieving specific capture and enrichment of modified nucleic acids. Firstly, this method efficiently enriched two major DNA modifications, 8-oxo-7,8-dihydroguanine (8-oxo-dG) and 5-hydroxymethylcytosine (5hmC) in model DNA systems. Then, genome-wide distribution of 8-oxodG from human cells and tissues align with previously reports. Finally, employing ‘Click-IP-Seq’, we performed the first comprehensive analysis of 8-oxo-dG spatial distribution and associated biological functions in human colorectal carcinoma tissues. This technology provides a high-specificity and versatile enrichment platform for nucleic acid modifications, which is expected to promote the application of cancer molecular diagnosis.
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