What is the main advantage of G4 CUT&Tag compared with G4 ChIP-seq?

G4 CUT&Tag profiles DNA G-quadruplexes directly in native chromatin using in situ tagmentation, which typically lowers background and reduces input requirements. This makes it more suitable for limited or precious samples while providing high-resolution G4 peak maps.

How much sample do I need for a G4 CUT&Tag project?

As a general guideline, we recommend around 1 × 10⁵ – 1 × 10⁶ viable cells or 20–50 mg tissue per sample. If your input is lower, share your estimated amounts and we will assess feasibility before starting.

Which species and sample types can you support?

We routinely work with human and mouse samples (cells and tissues) and can also support other mammalian species, model organisms, and selected non-mammalian systems on request. Both cultured cells and fresh/frozen research tissues are accepted.

Do I need controls and replicates for G4 CUT&Tag?

Yes. We generally recommend at least two biological replicates per condition and an appropriate negative control (e.g., IgG or no-primary-antibody CUT&Tag). We will help you choose controls and replicate numbers during study design.

How will I receive my data?

You receive raw FASTQ, aligned BAM, G4 peak and annotation files, visualisation-ready tracks, key plots, and a brief QC/summary report, all in analysis-friendly formats.

G4 CUT&Tag Service for Native DNA G-Quadruplex Mapping

Our G4 CUT&Tag service maps DNA G-quadruplexes (G4s) directly in native chromatin with high resolution and low background. Using BG4-based CUT&Tag chemistry, CD Genomics generates reliable G4 profiles from low-input samples for research use only.

Compared with G4 ChIP-seq and in vitro G4-seq, G4 CUT&Tag reduces input requirements and non-specific noise, helping you capture functional G4 sites involved in transcription, replication, DNA methylation, and genome stability.

BG4 G4 CUT&Tag chemistry targets pA-Tn5 to native DNA G-quadruplex structures, enabling precise genome-wide G4 mapping at promoters, enhancers, telomeres, and CpG islands.
Low-input CUT&Tag workflow delivers robust G4 profiles from as few as 1×10⁵–10⁶ cells, making G4 analysis feasible for rare cell populations, organoids, and precious research samples.
Integrated sequencing and analysis pipeline provides QC'd data, G4 peak calls, and functional annotation, so your team can move quickly from raw reads to interpretable G-quadruplex maps.

All CD Genomics sequencing services are for research use only.

Get a Quote for G4 CUT&Tag

Illustration of BG4-based G4 CUT&Tag sequencing workflow showing DNA G-quadruplex structures, BG4 antibody, pA-Tn5 tagmentation, and icons for low-input samples, high signal-to-noise, and integrated analysis.

Overview

What Is G4 CUT&Tag Sequencing

G4 CUT&Tag sequencing is a genome-wide method for mapping DNA G-quadruplex (G4) structures directly in native chromatin. It combines a G4-specific probe, such as the BG4 antibody, with the CUT&Tag workflow to generate high-resolution, low-background G4 profiles from low-input samples.

In a typical G4 CUT&Tag experiment, intact nuclei are immobilised on beads and incubated with a G4-binding antibody that recognises native G-quadruplexes on DNA. A secondary antibody then recruits a protein A–Tn5 transposase fusion, which inserts sequencing adapters next to antibody-bound G4 sites. After library preparation and high-throughput sequencing, the tagged fragments reveal the genomic distribution of DNA G-quadruplexes.

Key features at a glance

Native chromatin context – G4 structures are profiled in permeabilised nuclei, preserving the physiological chromatin environment instead of refolding DNA in vitro.
Low-input requirement – Optimized CUT&Tag chemistry enables genome-wide G4 mapping from relatively small numbers of cells or limited tissue.
High signal-to-noise – In situ tagmentation at antibody-bound sites reduces nonspecific background, improving peak detection at promoters, enhancers, telomeres, and other regulatory regions.

Applications

Why Map DNA G-Quadruplexes with G4 CUT&Tag

DNA G-quadruplexes (G4s) are four-stranded structures formed by guanine-rich sequences. They are enriched at promoters, enhancers, telomeres, CpG islands, and replication origins, where they can influence transcription, DNA replication, DNA methylation, and genome stability. G4 CUT&Tag sequencing lets you map these structures genome-wide in native chromatin, so you can distinguish functional G4s from simple sequence motifs.

G4 CUT&Tag is particularly useful when you want to:

Link G4s to gene expression by mapping G4 structures at key promoters and enhancers and comparing them with changes in mRNA levels, transcription factor binding, or histone marks.
Characterize regulatory G4s beyond promoters by profiling G4s in enhancers and UTRs and integrating the maps with ATAC-seq or RNA-seq to identify truly active regulatory sites.
Study replication, telomeres, and genome stability by locating G4s at replication origins and telomeres and assessing their association with fork stalling, telomere maintenance, or DNA damage.
Explore epigenetic cross-talk by combining G4 CUT&Tag with DNA methylation profiling (e.g., WGBS) to test how methylation patterns correlate with G4 formation around transcription start sites.
Support mechanism-of-action and target validation by tracking G4 landscape changes after helicase perturbation, oncogenic mutations, or treatment with G4-targeting compounds in cancer, neurodegeneration, infection, or developmental models.

By providing low-input, high-resolution DNA G-quadruplex maps in native chromatin, G4 CUT&Tag offers more mechanistic insight than prediction-only tools or bulk biochemical assays, while remaining practical for routine research projects.

Method Comparison

Choosing the Right G4 Mapping Method

Once you decide to study DNA G-quadruplexes, the key question is which mapping strategy best matches your sample type and research goal. The overview below compares G4 CUT&Tag sequencing with other commonly used G4 profiling approaches.

Attribute	G4 CUT&Tag	G4 ChIP-seq	In vitro G4-seq	G4 prediction tools
Sample input	Low–moderate cell numbers; limited tissue	High cell numbers; bulk tissue	Genomic DNA	No biological material
Context	Native chromatin in permeabilised nuclei	Crosslinked chromatin	DNA refolded in solution	In silico sequence analysis
Resolution & background	High resolution; low background	Moderate–high background; sonication noise	High motif resolution; no chromatin info	Motif-level only; no occupancy
Main strength	Native, condition-dependent G4 maps from low input	Established workflow in equipped labs	Detects potential G4-forming sequences	Rapid genome-wide G4 motif screening
Typical use case	Functional G4 profiling in limited or precious samples	Projects with abundant material and existing ChIP-seq infrastructure	Survey of G4 sequence propensity across the genome	Hypothesis generation and initial target selection

For projects that focus on native, condition-dependent G4 structures and have limited material, we generally recommend G4 CUT&Tag as the primary experimental approach. When needed, we can complement CUT&Tag data with in vitro G4-seq or computational prediction to explore sequence propensity and validate candidate regions in more detail.

Integrating G4 CUT&Tag with Other Omics

G4 CUT&Tag becomes most powerful when it is analysed alongside other genomic and epigenomic layers. CD Genomics can coordinate G4 CUT&Tag sequencing with complementary assays and deliver harmonised outputs for integrated analysis.

Typical multi-omics combinations

G4 CUT&Tag + ATAC-seq
Relate DNA G-quadruplexes to chromatin accessibility to see which G4 sites sit in open, regulatory regions.
G4 CUT&Tag + ChIP-seq / CUT&Tag
Overlay G4 maps with histone marks (e.g., H3K4me3, H3K27ac) or transcription factor binding (e.g., CTCF) to dissect regulatory mechanisms.
G4 CUT&Tag + DNA methylation (WGBS/RRBS)
Examine how DNA methylation patterns correlate with G4 formation around promoters and other regulatory elements.
G4 CUT&Tag + RNA-seq
Connect changes in G4 landscapes to gene expression profiles in the same samples or matched conditions.

Benefits of a coordinated design

Consistent sample handling and batch management across assays
Matched file formats and genome builds for straightforward integration
A single technical contact for planning, execution, and data delivery across the full multi-omics project

Advantages

Why Partner with CD Genomics for G4 CUT&Tag Sequencing

Selecting G4 CUT&Tag as a method is only the first step; execution quality and analysis depth determine how much insight you actually gain. CD Genomics combines optimized wet-lab workflows with dedicated bioinformatics to deliver consistent, publication-ready G4 CUT&Tag data.

Assay setup tailored to your system – We tune antibody conditions, nuclei preparation, and library parameters to your cell type, species, and sample quality, reducing trial-and-error and lowering the risk of failed libraries.

Consistent, review-ready data quality – Each project includes internal controls and standard QC metrics such as alignment rates, enrichment scores, and replicate concordance, so you can directly use the outputs in manuscripts and internal reports.

Built-in G4 CUT&Tag bioinformatics expertise – Our dedicated pipelines cover core G4 peak calling through to functional annotation and optional integration with other epigenomic or transcriptomic datasets, shortening the path from raw reads to biological conclusions.

Support for coordinated multi-omics designs – We can plan G4 CUT&Tag alongside ATAC-seq, ChIP-seq/CUT&Tag, WGBS/RRBS, RNA-seq, or Hi-C and deliver harmonized outputs, making it easier to compare G4 patterns with chromatin accessibility, histone marks, DNA methylation, or 3D genome structure.

Flexible project scopes from pilot to cohorts – Whether you are running a small feasibility study or a larger cohort, you work with a single technical contact who coordinates design, timelines, and deliverables across the entire project.

Together, these capabilities make CD Genomics a reliable partner for G4 CUT&Tag sequencing in discovery biology, target validation, and preclinical research.

Workflow

G4 CUT&Tag Workflow at CD Genomics

Our G4 CUT&Tag workflow efficiently takes you from samples to interpretable G-quadruplex maps.

Study design & sample review
We align on species, sample types, comparison groups, and input amounts, then recommend appropriate controls, replicates, and sequencing depth for your G4 CUT&Tag project.
Nuclei preparation & bead binding
Cells or tissues are gently processed to isolate intact nuclei, which are immobilised on ConA-coated magnetic beads to support antibody binding and in situ tagmentation.
G4 antibody binding in native chromatin
Bead-bound nuclei are incubated with a G4-specific antibody (BG4 or similar), followed by a secondary antibody to amplify signal and provide docking sites for the protein A–Tn5 (pA-Tn5) fusion.
pA-Tn5 tagmentation at G4 sites
pA-Tn5 is recruited to antibody-bound G4 structures and activated with magnesium, cutting DNA near G4 sites and inserting adapters directly in native chromatin.
Library preparation, sequencing & data handover
Adapter-tagged fragments are purified, amplified, QC'd, and sequenced on an Illumina platform. Cleaned reads and initial G4 peak calls are delivered and passed into the full bioinformatics pipeline described in the next section.

Vertical flowchart showing the CD Genomics G4 CUT&Tag workflow from study design and nuclei preparation through BG4 antibody binding, pA-Tn5 tagmentation, and Illumina sequencing to G-quadruplex peak maps.

Sample

Sample Requirements and Shipping Guidelines

To obtain high-quality G4 CUT&Tag sequencing data, we recommend planning samples with the following basic requirements in mind. All CD Genomics services are for research use only and not intended for diagnostic or therapeutic applications.

Supported sample types

Cultured cells: adherent or suspension cell lines, primary cells, organoids
Fresh or frozen tissues: animal or human research tissue, small research biopsies
Other models (yeast, bacteria, plant tissues, specialised samples): on consultation

Recommended input (guideline values)

Cultured cells: ~1 × 10⁵ – 1 × 10⁶ viable cells per sample (viability ideally ≥ 85–90%)
Tissues: ~20–50 mg per sample, depending on tissue type and cellularity

If your material is significantly below these ranges, share your estimated input; we will assess feasibility and adjust expectations before starting.

Collection and storage

Avoid Trizol or other strong lysis/extraction reagents for G4 CUT&Tag samples.
Cells: harvest gently, freeze in appropriate freezing medium, store in liquid nitrogen or at −80 °C.
Tissues: snap-freeze as soon as possible and store at −80 °C in clearly labelled tubes.

Shipping

Ship samples on dry ice in insulated containers.
Avoid repeated freeze–thaw cycles; plan a single shipment with sufficient dry ice.
Include a simple sample list (IDs, type, treatment, estimated input) matching your digital submission.

Sequencing

Sequencing Strategy and Recommended Depth

Our G4 CUT&Tag sequencing service is optimized to balance resolution, cost, and downstream analysis requirements for genome-wide DNA G-quadruplex mapping.

Platform and read configuration

Illumina short-read platforms (e.g., NovaSeq or equivalent)
Typical configuration: paired-end 2 × 50 bp or 2 × 75 bp, suitable for CUT&Tag fragment sizes and peak calling
Other read lengths can be arranged for specific study needs

Recommended sequencing depth

As a general guide for bulk G4 CUT&Tag:

Standard G4 profiling:

~20–30 million uniquely mapped reads per sample for robust peak calling across the genome

Higher-resolution or complex genomes:

Deeper coverage can be requested if you need finer peak definition or plan extensive integrative analyses

We will tailor depth recommendations based on:

Genome size and species
Number of conditions and replicates
Whether you plan to integrate G4 CUT&Tag with ATAC-seq, ChIP-seq, WGBS, RNA-seq, or Hi-C

Controls and replicates

To support reliable interpretation, we generally recommend:

At least 2 biological replicates per condition
An appropriate negative control (e.g., IgG or no-primary-antibody CUT&Tag)
Optional additional controls if you are testing G4-targeting ligands, helicase perturbations, or other interventions

Our team will help you choose a sequencing plan that matches your experimental question, ensuring that G4 CUT&Tag data are generated at a depth that supports confident DNA G-quadruplex peak detection and downstream analysis.

Bioinformatics

G4 CUT&Tag Data Analysis

Our G4 CUT&Tag data analysis pipeline is built to turn raw sequencing reads into reliable, interpretable DNA G-quadruplex maps, without the need for in-house bioinformatics scripting.

Vertical flowchart of the G4 CUT&Tag bioinformatics analysis workflow, from raw FASTQ QC and alignment through G-quadruplex peak calling, quality control, peak annotation, differential and motif analysis, to GO/KEGG enrichment and reporting.

1. Primary processing

Demultiplexing and adapter trimming
Read quality filtering and alignment to the reference genome
Mapping statistics to summarise alignment rates and coverage

2. G4 peak calling and quality control

Peak calling optimized for G4 CUT&Tag signal profiles
Global QC metrics, including enrichment scores and background assessment
Replicate concordance checks to highlight robust, reproducible G4 peaks

3. Functional and genomic annotation

Annotation of G4 peaks relative to promoters, enhancers, UTRs, exons, introns, and intergenic regions
Linking G4 sites to nearby genes and transcription start sites
Optional GO / pathway-style enrichment for genes associated with G4-enriched regions

4. Integrative analysis (optional)

On request, we can integrate G4 CUT&Tag data with:

Chromatin accessibility (ATAC-seq)
Histone marks or transcription factor binding (ChIP-seq / CUT&Tag)
DNA methylation profiles (e.g., WGBS)
Gene expression data (RNA-seq)

This places DNA G-quadruplex structures in the broader context of gene regulation and chromatin state, supporting clearer mechanistic conclusions.

Deliverables

G4 CUT&Tag Deliverables and Demo Results

We provide a concise, analysis-ready output package so your team can directly use the data in downstream work.

Data files

Raw FASTQ files for all G4 CUT&Tag libraries and controls
Aligned BAM files + index files mapped to the agreed reference genome

Peak and annotation outputs

G4 peak files (e.g., BED format) for each sample or condition
Annotation tables linking peaks to genomic features and nearest genes/TSS

Visuals and report

Genome browser tracks and key plots (e.g., peak distribution, TSS enrichment, sample correlations)
A brief QC and summary report describing methods, main quality metrics, and example G4 maps

GO enrichment bubble plot showing biological process terms, where bubble size reflects gene count and colour indicates enrichment score for G4 CUT&Tag–associated genes. GO enrichment bubble plot

Bar plot showing chromosome lengths with red bands marking G4 CUT&Tag peak density across each chromosome. Chromosome-level distribution

TSS-centred heatmaps with line plots showing signal intensity across multiple samples Heatmaps and average profiles

PCA scatter plot of G4 CUT&Tag samples, with red control replicates separated from clustered blue treated replicates along PC1 and PC2. PCA plot

Case

Case Study

Genome-wide G4 CUT&Tag Mapping in Mouse Embryonic Stem Cells

Genome-wide mapping of G-quadruplex structures with CUT&Tag. Nucleic Acids Research.

Lyu J, Shao R, Yung PYK, Elsässer SJ., Nucleic acids research, 2022.

1. Background

Predicted G-quadruplex motifs are widespread in the genome, but only some form stable structures on chromatin. Lyu et al. set out to use G4 CUT&Tag to map endogenous DNA G-quadruplexes in mouse embryonic stem cells (ESCs) and to define where chromatin G4s actually occur.

2. Methods

The authors combined FLAG-tagged BG4 antibody with CUT&Tag in permeabilized ESC nuclei, using protein A–Tn5 to tagment DNA adjacent to BG4-bound G4s and calling peaks genome-wide. They further applied a quantitative G4 CUT&Tag strategy with spike-in DNA, nuclease treatment to test single-stranded dependence, and integration with R-loop and other epigenomic datasets.

3. Results

G4 CUT&Tag identified high-confidence G4 peaks enriched at active promoters and enhancers, with low signal in gene bodies and repressed regions.
Active promoters often carried canonical G4 motifs, while active/poised enhancers were enriched for non-canonical G4 motifs, distinguishing them from primed enhancers lacking G4 motifs and signal.
Upon differentiation from ESCs to neural progenitors, many enhancer G4s were lost in parallel with gene expression changes.
Nuclease and R-loop mapping confirmed that G4 CUT&Tag signal depends on locally melted DNA and frequently co-occurs with R-loops at regulatory elements.

4. Conclusions

This work shows that G4 CUT&Tag is a sensitive, native chromatin method for mapping DNA G-quadruplexes and that G4s are characteristic of active promoters and enhancers in ESCs. It illustrates how G4 CUT&Tag, combined with other assays, can reveal how G4 structures mark functional regulatory elements and change across cell states—exactly the type of insight G4 CUT&Tag sequencing can provide in research projects.

Comparison of G4 CUT&Tag to G4 ChIP-seq and G4P-ChIP.

FAQ

FAQs: G4 CUT&Tag Sequencing Service

References

Lyu, J., Shao, R., Yung, P.Y.K. et al. Genome-wide mapping of G-quadruplex structures with CUT&Tag. Nucleic Acids Research 50, e13 (2022).
Kaya-Okur, H.S., Wu, S.J., Codomo, C.A. et al. CUT&Tag for efficient epigenomic profiling of small samples and single cells. Nature Communications 10, 1930 (2019).
Chambers, V.S., Marsico, G., Boutell, J.M. et al. High-throughput sequencing of DNA G-quadruplex structures in the human genome. Nature Biotechnology 33, 877–881 (2015).
Johnson, S.A., Paul, T., Sanford, S.L. et al. BG4 antibody can recognize telomeric G-quadruplexes harboring destabilizing base modifications and lesions. Nucleic Acids Research 52, 1763–1778 (2024).
Varshney, D., Spiegel, J., Zyner, K.G. et al. The regulation and functions of DNA and RNA G-quadruplexes. Nature Reviews Molecular Cell Biology 21, 459–474 (2020).
Georgakopoulos-Soares, I., Chan, C.S.Y., Ahituv, N. et al. High-throughput techniques enable advances in the roles of DNA and RNA secondary structures in transcriptional and post-transcriptional gene regulation. Genome Biology 23, 159 (2022).
Qian, S.H., Shi, M.W., Xiong, Y.L. et al. EndoQuad: a comprehensive genome-wide experimentally validated endogenous G-quadruplex database. Nucleic Acids Research 52, D72–D80 (2024).