Mapping the complete spectrum of DNA modifications requires tools that move beyond conventional methylation analysis to capture the dynamic process of active demethylation. To enable sensitive mapping of the whole genomes of key oxidation intermediates 5fC and 5caC, CD BioSciences offers M.SssI methyltransferase-assisted bisulfite sequencing (MAB-seq) services, thereby advancing your research in neurobiology, developmental biology, and disease mechanisms.
M.SssI methylase-assisted bisulfite sequencing (MAB-seq) is a specialized enzymatic-chemical method designed for the sensitive, genome-wide mapping of the rare cytosine oxidation products 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC). The technique employs a two-step protection strategy: first, M.SssI methyltransferase treatment adds a methyl group to all unmodified cytosines (C) at CpG sites, converting them to 5-methylcytosine (5mC) and thereby "protecting" them. Subsequent bisulfite conversion then specifically deaminates the unprotected 5fC and 5caC to uracil (U), which is read as thymine (T) during sequencing. Consequently, in the final sequence data, only the original 5fC and 5caC sites appear as T, while the protected 5mC remain as C, enabling their unambiguous identification and quantification against a background of pre-existing and enzymatically generated 5mC.
Fig.1 The common bisulfite-conversion-based techniques to analyze DNA methylation in human cancers. (Jeddi F, et al., 2024)
M.SssI methylase-assisted bisulfite sequencing (MAB-seq) offers a powerful and refined approach for probing the intricate landscape of active DNA demethylation. By employing an elegant enzymatic protection strategy, it overcomes the significant challenges of detecting the rare and transient oxidative bases 5fC and 5caC, providing researchers with a direct, high-resolution, and quantitative tool to investigate these critical epigenetic intermediates.
Direct & Simultaneous Detection: Enables the direct, concurrent mapping of both 5fC and 5caC in a single experiment without complex data subtraction.
Single-Base Resolution: Provides precise, nucleotide-level localization of these rare oxidative modifications across the genome.
Quantitative Capability: Delivers data on the relative abundance of 5fC and 5caC at each site, allowing for comparative analysis.
Streamlined & Cost-Effective: Offers a more efficient and economical workflow compared to older subtraction-based methods, especially when using targeted approaches like RR-MAB-seq.
As a leading provider of advanced epigenetic research services, CD BioSciences offers expert M.SssI methylase-assisted bisulfite sequencing (MAB-seq) to empower your investigation of active DNA demethylation dynamics. Our service is distinguished by a rigorously optimized enzymatic protection step and ultra-high bisulfite conversion efficiency, ensuring the sensitive and accurate genome-wide profiling of the rare oxidative marks 5fC and 5caC.
Our MAB-seq service follows a meticulously optimized, end-to-end pipeline specifically designed to preserve the integrity of the rare 5fC and 5caC modifications while ensuring maximum enzymatic conversion efficiency and sequencing data quality. From initial sample assessment to final data interpretation, each step is controlled and validated to deliver reliable, high-resolution maps of these key oxidative bases.
Project Consultation & Sample QC
We begin by collaborating with you to confirm the experimental goals and assess the suitability of your genomic DNA. Rigorous quality control is performed to ensure sufficient quantity, purity, and integrity for optimal MAB-seq library construction.
M.SssI Methyltransferase Treatment (Enzymatic Protection)
This is the critical and differentiating step. Your DNA is treated with the M.SssI methyltransferase under optimized conditions to efficiently convert all unmodified cytosines (C) to 5-methylcytosine (5mC), thereby "protecting" the background and leaving the target 5fC and 5caC residues unaltered and exposed.
Bisulfite Conversion & Library Preparation
The M.SssI-treated DNA undergoes high-efficiency bisulfite conversion, which specifically deaminates the unprotected 5fC and 5caC to uracil. The converted DNA is then purified and constructed into a high-quality, sequencing-ready library.
High-Throughput Sequencing & Primary Analysis
Libraries are sequenced on Illumina platforms to achieve the required depth. The raw data is processed through primary analysis, including basecalling, demultiplexing, and alignment to a bisulfite-converted reference genome.
Dedicated MAB-seq Data Analysis & Reporting
We employ specialized bioinformatics pipelines to call 5fC/5caC sites from the unique T-signals in the aligned data. We deliver a comprehensive report including genome-wide maps, quantitative analysis, annotation of sites relative to genomic features, and expert interpretation support.
We provide a complete, start-to-finish service. You can submit the biological samples directly, and our experts will handle the entire process, from nucleic acid extraction to final data analysis. The sample types compatible with our optimized workflow include, but are not limited to:
CD BioSciences provides comprehensive solutions for advanced DNA epigenetic analysis, empowering researchers to decode complex regulatory layers with precision and confidence. Contact us to navigate the full spectrum of DNA modifications and accelerate your discovery in development, disease, and beyond.
Reference 1. Jeddi F, Faghfuri E, Mehranfar S, et al. The common bisulfite-conversion-based techniques to analyze DNA methylation in human cancers[J]. Cancer Cell International, 2024, 24(1): 240.
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