Once considered rare in eukaryotes, DNA N6-methyladenine (6mA) is now recognized as a dynamic and functional epigenetic mark in mammals, involved in transcriptional regulation, transposable element silencing, and cellular stress response. CD BioSciences offers precise, sensitive, and reliable 6mA detection and mapping services to help researchers explore its emerging roles in development, disease, and environmental adaptation.

Introduction to N6-Methyladenine (6mA)

N6-Methyladenine (6mA) is an epigenetic modification where a methyl group is added to the sixth nitrogen of the adenine base. While historically studied in bacteria and plants, 6mA has gained significant attention in recent years as a potentially regulatory mark in mammalian and other eukaryotic systems. It is catalyzed by methyltransferases such as METTL4 and N6AMT1 and can be actively removed by demethylases like ALKBH1. Functionally, 6mA in eukaryotes has been associated with diverse processes, including transcriptional activation, repression of transposable elements, regulation of chromatin structure, and cellular response to environmental stressors. Unlike 5mC, which is enriched in symmetrical CpG contexts, 6mA often occurs in non-CpG motifs, suggesting distinct regulatory logic. Dysregulation of 6mA has been implicated in cancer progression, neurological disorders, and developmental defects, making it a promising new frontier for epigenetic biomarker discovery and therapeutic targeting.

Fig.1 Illustration of N6-methyladenine (6mA) modifications in DNA. (Lv H, et al., 2019)

Our Services

With a deep understanding of the technical complexities involved in detecting low-abundance epigenetic marks, CD BioSciences delivers end-to-end solutions for the sensitive and specific analysis of DNA N6-methyladenine (6mA). By integrating validated workflows that combine antibody-based enrichment, third-generation sequencing, and mass spectrometry, we provide high-confidence 6mA profiles designed to accelerate your research in novel epigenetic regulatory mechanisms.

6mA Analysis Solutions by CD BioSciences

We employ a multi-platform strategy to ensure accurate 6mA detection, tailored to your resolution and throughput needs.

1. Global DNA Modification Quantification by LC-MS/MS: Provides absolute quantification of 6mA with gold-standard accuracy and specificity, ideal for validating global modification levels.
2. DNA Immunoprecipitation (DIP) Sequencing: Utilizes validated high-specificity anti-6mA antibodies for immunoprecipitation and sequencing to deliver genome-wide 6mA distribution profiles.
3. Nanopore Sequencing: Enables direct, real-time detection of 6mA on long native DNA strands without amplification or chemical conversion.
4. PacBio SMRT DNA Sequencing: Achieves high-confidence 6mA mapping through kinetic analysis during single-molecule, real-time sequencing.

Workflow of DNA 6mA Analysis Service

High-Integrity Genomic DNA Preparation

We perform stringent DNA extraction and quality assessment, including fluorometric quantification, spectrophotometric purity analysis, and fragment size profiling, to ensure input material meets the exacting requirements of downstream 6mA-sensitive protocols.

Platform-Specific 6mA Processing

1. For IP-based methods: DNA undergoes immunoprecipitation with rigorously validated anti-6mA antibodies under optimized buffer conditions.
2. For SMRT/Nanopore sequencing: DNA is prepared as high-molecular-weight libraries without disruptive bisulfite conversion, preserving native modification states.
3. For LC-MS/MS: DNA is enzymatically digested to nucleosides for direct instrumental analysis.

Controlled Library Construction and Sample Loading

We prepare sequencing libraries with minimal PCR cycles to reduce bias or prepare purified digests for LC-MS/MS. Each batch includes internal controls and spike-ins to monitor enrichment efficiency, sequencing kinetics, or instrument performance.

High-Resolution Data Acquisition and Primary Processing

Sequencing is performed on Illumina (for IP-seq), PacBio Revio/Sequel IIe, or Oxford Nanopore PromethION platforms, while quantification utilizes high-sensitivity triple-quadrupole LC-MS/MS. Raw data is processed through platform-specific primary pipelines (e.g., IP peak calling, kinetic variant detection).

Advanced Bioinformatic Analysis and Epigenomic Interpretation

Our custom pipelines perform differential 6mA analysis, genomic context annotation (promoters, enhancers, repeats), and correlation with orthogonal datasets (e.g., RNA-seq, ATAC-seq). We utilize established tools (e.g., MetaGene, Nanopolish) and proprietary algorithms to distinguish true modification signals from noise.

Our Advantages

1. Multi-Platform Validation: We cross-validate findings using complementary technologies (e.g., IP-seq with LC-MS/MS) to ensure data reliability.
2. Expertise in Low-Abundance Modification Detection: Our protocols are optimized for the sensitive and specific capture of 6mA signals.
3. Single-Molecule Resolution with Long Reads: Third-generation sequencing provides haplotype-phased 6mA information across complex genomic regions.
4. Integrated Functional Analysis: Our bioinformatics team correlates 6mA patterns with gene expression and chromatin data to uncover regulatory insights.
5. Rigorous Quality Control: Every step, from antibody validation to final data analysis, follows stringent QC protocols.

CD BioSciences is your dedicated partner for comprehensive DNA modification analysis. In addition to 6mA, we provide full-service solutions for 5mC, 5hmC, 5fC, 5caC, 4mC, and 8-oxoG. Our flexible, project-tailored approach ensures you receive the most appropriate technology and deepest biological insight for your research in development, disease, or environmental epigenetics. Contact us to design your customized epigenetic study.

Reference
1. Lv H, Dao F Y, Guan Z X, et al. iDNA6mA-Rice: a computational tool for detecting N6-methyladenine sites in rice[J]. Frontiers in genetics, 2019, 10: 793.