5fC Chemically Assisted Bisulfite Sequencing (fCAB-seq) Service

Conventional bisulfite sequencing (BS-seq) faces a significant challenge in distinguishing 5-formylcytosine (5fC) from other cytosine modifications like 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC), leading to the masking of critical 5fC signals. To address this, CD BioSciences offers a state-of-the-art 5fC chemically assisted bisulfite sequencing (fCAB-seq) service, empowering researchers to accurately decode the dynamic landscape of active DNA demethylation.

Introduction to fCAB-seq

Traditional BS-seq treats 5mC, 5hmC, and 5fC identically, converting unmodified cytosine to uracil while leaving these three modifications read as cytosine. This inability to discriminate between them obscures the unique biological signal of 5fC, an important oxidative intermediate in active DNA demethylation linked to gene regulation and development.

fCAB-seq overcomes this limitation through a clever chemical intervention prior to bisulfite conversion. The core principle involves the selective chemical reduction of 5fC to 5hmC using a reducing agent like sodium borohydride (NaBH4). In a subsequent standard BS-seq reaction, the original 5hmC residues are read as C, while the "newly reduced 5fC-derived 5hmC" and the original 5mC are read as T. By bioinformatically comparing the sequencing results from fCAB-seq with those from conventional BS-seq or other methods (like oxBS-seq for 5hmC), 5fC sites can be uniquely identified at single-base resolution.

Fig.1 Principal scheme of oxBS-seq and fCAB-seq methods. (Becker D, et al., 2014)

Features of fCAB-seq

The fCAB-seq technology itself is distinguished by several key features that make it a powerful tool for epigenetic research, addressing specific limitations of conventional bisulfite sequencing. These inherent characteristics enable precise investigation into the role of 5fC.

1. High Specificity: Utilizes selective chemical reduction to uniquely target 5fC, effectively distinguishing it from 5mC, 5hmC, and 5caC.
2. Single-Base Resolution: Enables nucleotide-level mapping of 5fC sites across the entire genome for precise localization.
3. Quantitative Capability: Allows direct comparison of 5fC abundance at specific loci between different samples or conditions.
4. Genome-Wide Coverage: Provides an unbiased and comprehensive profile of 5fC distribution in all genomic contexts.
5. Chemical Consistency: Employs a defined chemical reaction, enhancing experimental reproducibility and reducing variability.

Our Services

CD BioSciences provides a complete, end-to-end fCAB-seq service for the detection and analysis of 5-formylcytosine (5fC). We help our clients overcome technical hurdles and obtain high-quality, publication-ready data. Our service encompasses expert experimental execution, robust bioinformatics analysis, and detailed reporting, accelerating your research into DNA demethylation dynamics, cellular differentiation, and disease mechanisms.

Workflow of fCAB-seq Service

At CD BioSciences, we ensure a seamless and expertly managed journey from your initial inquiry to final report delivery. Our established, end-to-end workflow is meticulously designed to deliver high-quality, reliable data while maximizing project efficiency and scientific value.

Consultation & Project Design

Our service begins with a collaborative consultation. Our expert scientists will review your research objectives and provide a customized project plan, including sample requirements, sequencing depth recommendations, and a tailored bioinformatics analysis strategy to ensure your specific hypotheses are addressed.

Sample Submission

Upon project confirmation, you will receive detailed guidelines for sample preparation and shipment. Our lab accepts a variety of sample types, including purified genomic DNA, cells, and tissues. Rigorous quality control is performed upon receipt to verify integrity and quantity before proceeding.

Library Preparation & Sequencing

This core phase is executed by our specialized technical team. The process includes the selective chemical reduction of 5fC, followed by bisulfite conversion, high-efficiency library construction, and high-throughput sequencing on Illumina platforms. Strict QC checkpoints are implemented at every step to ensure optimal library quality and data yield.

Data Analysis

Your sequencing data is processed through our validated, proprietary bioinformatics pipeline. Our analysis includes precise alignment, 5fC site calling, genome annotation, differential modification analysis, and the generation of comprehensive visualizations. Advanced, custom integration with other omics datasets is available upon request.

Report Delivery

You will receive a complete, interpretable project report. The final delivery includes all raw and processed data, publication-ready figures and charts, a detailed methodological description suitable for publications, and a clear summary of the key biological findings and insights.

Supported Sample Types

1. Purified Genomic DNA
2. Cultured Cells (Mammalian, etc.)
3. Fresh-Frozen Tissue

1. Blood and Blood Products
2. FFPE Tissue Sections
3. Other Types (Please Inquire)

Our Advantages

1. Expert-Optimized Protocol: Leveraging deep epigenetic expertise and a rigorously validated fCAB-seq workflow for highest data reliability.
2. Complete Turnkey Solution: Seamless, end-to-end service from project design to final interpretation, managed by dedicated specialists.
3. Advanced Custom Bioinformatics: Delivering deep insights through powerful, customizable analysis and optional multi-omics data integration.
4. Rigorous Quality Assurance: Implementing strict QC checkpoints at every stage to ensure robust, publication-ready results.
5. Collaborative & Secure Partnership: Providing direct scientist support and guaranteed data confidentiality throughout the project.

To further support comprehensive 5fC research, CD BioSciences also provides a suite of complementary services, including global DNA modification quantification by LC-MS/MS and ELISA, as well as advanced sequencing solutions such as CLEVER-seq and MAB-seq. If you are interested in our services, please feel free to contact us for more details and quotation information of related services.

Reference 1. Becker D, Lutsik P, Ebert P, et al. BiQ Analyzer HiMod: an interactive software tool for high-throughput locus-specific analysis of 5-methylcytosine and its oxidized derivatives[J]. Nucleic acids research, 2014, 42(W1): W501-W507.