What is G-loop analysis and why is it important?

G-loop analysis identifies genomic regions where G-quadruplexes (G4s) and R-loops overlap, forming three-stranded nucleic acid structures known as G-loops. These regions play a vital role in transcription regulation, replication stress response, and genome stability. Understanding their distribution helps researchers reveal how RNA–DNA interactions shape gene expression and chromatin architecture.

How does G-loop Analysis differ from ChIP-seq or DRIP-seq?

Unlike ChIP-seq or DRIP-seq, G-loop Analysis uses Cut&Tag chemistry, which performs in situ DNA tagging without crosslinking or sonication. This preserves native G4 and R-loop conformations and produces data with higher resolution, lower background noise, and improved reproducibility. It also requires far fewer cells than traditional immunoprecipitation-based methods.

What kind of samples are suitable for G-loop Analysis?

G-loop Analysis accepts a wide range of sample types, including cultured cells, animal tissues, and plant tissues. The key requirement is that the samples are viable, properly preserved, and from species with available reference genomes. Maintaining chromatin integrity is essential for reliable detection of G4 and R-loop structures.

What information can researchers obtain from G-loop sequencing data?

Researchers receive genome-wide maps of G4 and R-loop signals and integrated G-loop hotspot regions, along with associated motif analyses, GO and KEGG enrichment, and annotated gene lists. These results reveal how G4 and R-loop interactions influence transcriptional regulation, DNA repair pathways, and genome structure.

Is G-loop Analysis applicable to disease or drug research?

Yes. G-loop mapping supports the investigation of diseases involving transcriptional dysregulation or genome instability, such as cancer and neurodegenerative disorders. It also provides valuable insights for drug discovery, particularly for developing small molecules targeting G-quadruplex structures or molecular glue degraders that recognize G-loop motifs.

Can G-loop Analysis be integrated with other omics data?

Absolutely. Results from G-loop Analysis can be integrated with RNA-seq, ATAC-seq, or ChIP-seq datasets to correlate structural features with gene expression, chromatin accessibility, and histone modifications, providing a multi-dimensional view of genome function.

What support does CD Genomics provide after data delivery?

Every project includes detailed bioinformatics reports, visual figures, and expert guidance for data interpretation. Clients receive publication-ready outputs and have the option to discuss results with our scientific team to align findings with their experimental goals.

RNA Interaction
G-loop Analysis (G4 & R-loop Cut&Tag)

G-loop Analysis (G-quadruplexes & R-loop Cut&Tag) Service

Uncover the hidden genome-structure layer that governs transcription and genome stability. At CD Genomics, our G-loop Analysis Service brings together the power of G-quadruplexes (G4s) profiling and R-loop Cut&Tag to precisely map G-loop assembly across the genome. This hybrid approach empowers biotech and pharma researchers, CROs, and academic institutions to detect co-localised G4–R-loop regions with high resolution, minimal input and robust reproducibility. Gain actionable insight into chromatin architecture, replication stress, and non-coding RNA regulation.

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G-loop Analysis showing G-quadruplex (G4) and R-loop Cut&Tag integration for genome-wide G-loop hotspot mapping.

Genome-wide mapping of G-quadruplexes (G4s) and R-loops
High-resolution detection of G-loop assembly sites
Ultra-low sample input; high signal-to-noise ratio
Ideal for transcriptional regulation, epigenetics & drug-discovery teams

Overview Principle Workflow Analysis Advantages Applications Why Us Samples Results FAQs Inquiry

Understanding G-loop Analysis (G-quadruplexes & R-loop Cut&Tag)

G-loops represent a newly characterized three-stranded nucleic acid structure formed when G-quadruplexes (G4s) and R-loops coexist at the same genomic region. These dynamic structures play a central role in regulating transcription, replication, and genome stability. However, traditional sequencing methods often fail to distinguish G4–R-loop interactions or detect G-loop assembly in situ.

CD Genomics' G-loop Analysis Service integrates G4 Cut&Tag and R-loop Cut&Tag sequencing into a unified workflow. By combining the high specificity of BG4 (G4) and S9.6 (R-loop) antibodies with the precision of Tn5-based Cut&Tag chemistry, this platform enables direct, high-resolution identification of G-loop regions across the genome without disrupting native chromatin structures.

This service provides researchers with a powerful tool to:

Explore how G4s and R-loops coordinate transcriptional regulation.
Investigate genomic instability and replication stress mechanisms.
Support drug discovery, including small-molecule screening targeting G4–R-loop dynamics.

Recommended reading: Learn more about related methods such as R-loop Cut&Tag sequencing and DRIPc-seq, which complement G-loop analysis in understanding RNA–DNA hybrid formation.

Technology Principle

Cut&Tag-Based Profiling of G-quadruplexes and R-loops

The G-loop Analysis Service is built upon the Cut&Tag (Cleavage Under Targets and Tagmentation) platform — an advanced chromatin profiling method that enables in situ detection of protein–DNA or nucleic acid–DNA interactions without crosslinking or sonication. This approach preserves the native state of G4 and R-loop structures, ensuring authentic signal capture.

Dual Antibody Strategy

BG4 antibody: Recognizes G-quadruplex (G4) structures that form within guanine-rich DNA regions.
S9.6 antibody: Specifically binds to RNA–DNA hybrids (R-loops) that arise during transcription.

Using ProteinA–Tn5 transposase fusion enzymes, both antibodies guide Tn5 to their target regions, simultaneously cleaving DNA and inserting sequencing adapters. When G4 and R-loop signals overlap, the resulting intersection defines G-loop hotspots—regions where G4 folding and R-loop formation co-occur.

Why Cut&Tag for G-loop Analysis?

Preserves chromatin integrity — no harsh fragmentation steps.
High signal-to-noise ratio — ideal for low-abundance structures.
Requires fewer cells than ChIP-seq or DRIP-seq methods.
Delivers high-resolution peak data for precise genomic mapping.

This combined strategy not only maps individual G4 and R-loop landscapes but also identifies their co-localization, uncovering structural interactions crucial for transcriptional regulation and genomic stability.

Recommended reading:

To explore these complementary approaches, visit R-loop sequencing (DRIP-seq) or R-loop Cut&Tag sequencing.

Principle of G-loop Analysis using dual G-quadruplex (G4) and R-loop Cut&Tag assays for G-loop hotspot mapping. Dual Cut&Tag profiling with BG4 and S9.6 antibodies enables precise genome-wide identification of G-loop hotspots, preserving native chromatin integrity and delivering high-resolution mapping data.

Comprehensive Workflow for G-loop Analysis (G4 & R-loop Cut&Tag Integration)

CD Genomics provides a fully integrated workflow for G-loop Analysis, combining experimental precision with robust bioinformatics. Each step—from sample preparation to hotspot identification—is optimized to preserve native G-quadruplex (G4) and R-loop structures and ensure reproducible, high-resolution data output.

Step 1. Sample Preparation

Fresh cells or tissues are collected and converted into cell suspensions. No crosslinking or harsh fragmentation is required, ensuring the preservation of in vivo G4 and R-loop configurations.

Step 2. Antibody Binding

BG4 antibody specifically recognizes G4 structures.
S9.6 antibody targets RNA–DNA hybrids (R-loops).
Each antibody is incubated sequentially with the sample to label the respective targets.

Step 3. Protein A–Tn5 Recruitment & Tagging

The Protein A–Tn5 transposase complex binds to the antibody–target complex and performs site-specific DNA cleavage and adapter insertion, directly producing sequencing-ready fragments.

Step 4. Library Construction & Sequencing

Fragmented DNA is purified and amplified for high-throughput sequencing using Illumina or Nanopore platforms, providing genome-wide coverage of G4 and R-loop signals.

Step 5. Bioinformatics Analysis

Data are quality-controlled, aligned to the reference genome, and processed with MACS2 to identify enriched peaks. Overlapping G4 and R-loop peaks are merged to define G-loop hotspots, followed by functional annotation and enrichment analyses (GO, KEGG, and motif discovery).

Step 6. Reporting & Interpretation

Clients receive detailed reports including chromosomal distribution, differential peak analysis, motif enrichment, and GO/KEGG results—ready for publication or downstream research.

Bioinformatics Analysis

CD Genomics provides a comprehensive bioinformatics analysis pipeline for G-loop Analysis (G4 & R-loop Cut&Tag).

Our services are divided into Basic Analysis and Advanced Analysis, designed to meet the needs of both exploratory and in-depth studies.

Analysis Type	Content	Deliverables
Basic Analysis	Raw data quality control (QC) Adapter trimming and removal of low-quality reads Read alignment to reference genome (BWA or Bowtie2) Peak calling using MACS2 for G4 and R-loop signals Peak annotation and basic visualization	Clean FASTQ files Alignment (BAM) files Peak list (BED format) Annotated gene list Summary QC report (mapping rate, duplication rate, signal distribution)
Advanced Analysis	Differential peak analysis between experimental groups G4–R-loop co-localization and G-loop hotspot identification Motif enrichment analysis for binding site prediction Gene Ontology (GO) and KEGG pathway enrichment Correlation heatmap and clustering analysis Chromosome-level visualization and integrated report	Differential peak table G-loop hotspot map Motif logo plots GO/KEGG enrichment charts Correlation heatmap Comprehensive bioinformatics report (PDF + figures)

Software & Tools:

MACS2 • HOMER • BEDTools • deepTools • ChIPseeker • ClusterProfiler • RStudio visualization suite

Data Format Delivered:

FASTQ, BAM, BED, XLSX/CSV, PDF, PNG/SVG (publication-ready figures)

End-to-End Workflow of G-loop Analysis.

Technical Advantages

CD Genomics' G-loop Analysis (G4 & R-loop Cut&Tag) platform is designed for high precision, low noise, and exceptional reproducibility. This integrative method captures genome-wide G-loop hotspots under near-physiological conditions, setting a new benchmark for nucleic acid structure profiling.

1. Low Input, High Sensitivity

The optimized Cut&Tag chemistry enables successful profiling with as few as 2×10⁵ cells, making it ideal for rare or limited samples. Despite low input, the method delivers high signal intensity and clear G-loop peak definition.

2. Preserves Native Chromatin State

Unlike traditional ChIP-seq or DRIP-seq, this workflow avoids crosslinking and sonication. It maintains natural G4 and R-loop conformations, allowing accurate mapping of structures as they occur in vivo.

3. High Resolution and Low Background

Precise Tn5 transposase activity produces sharp, high-resolution peaks with minimal background noise. The result is a clearer, more confident identification of true G-loop regions.

4. Excellent Reproducibility

Optimized protocols and bioinformatics pipelines ensure strong data consistency across replicates and experimental batches—essential for comparative genomic studies.

Applications

1. Transcriptional Regulation Studies

G-loops serve as key regulators of transcription initiation and elongation. Mapping their locations helps researchers uncover how RNA transcripts modulate chromatin accessibility and gene activation.

2. Genome Stability and Replication Stress Research

Persistent G4 or R-loop structures can interfere with DNA replication and repair. G-loop analysis enables identification of regions prone to replication stress, aiding studies on genome instability and DNA damage responses.

3. Epigenetic and Chromatin Dynamics

Integrating G-loop data with histone modification or Hi-C datasets reveals how three-dimensional chromatin architecture influences gene expression. This supports multi-omics investigations into chromatin remodeling.

4. Disease Mechanism Exploration

Aberrant G4 or R-loop formation has been linked to cancer, neurodegenerative disorders, and aging. G-loop mapping provides insights into how dysregulated RNA–DNA interactions contribute to disease pathology.

5. Drug Discovery and Target Validation

Recent Science studies demonstrate that protein-binding "G-loop motifs" can influence drug–target interactions. Combining G-loop data with compound screening offers new perspectives for molecular glue and G4-targeted therapeutic development.

Why Choose CD Genomics

CD Genomics combines technical expertise, advanced sequencing platforms, and comprehensive bioinformatics to deliver reliable results for complex genome structure analysis. Our G-loop Analysis (G4 & R-loop Cut&Tag) service offers a complete solution from experimental design to data interpretation—ideal for researchers seeking reproducibility, depth, and accuracy.

1. Proven Expertise in Chromatin and RNA–DNA Structure Profiling

Our team has extensive experience in Cut&Tag, DRIP-seq, and R-loop mapping technologies. We have optimized G4 and R-loop detection workflows to capture native structures with minimal artifacts.

2. End-to-End Service Workflow

From sample preparation to final report delivery, every step is handled by our in-house experts. Clients receive validated data and professional analysis support without the need for additional downstream processing.

3. Integrated Multi-Omics Support

CD Genomics provides flexible integration with RNA-seq, ATAC-seq, or ChIP-seq data, enabling clients to correlate G-loop formation with gene expression, chromatin accessibility, and epigenetic modifications.

4. Quality, Transparency, and Publication-Ready Output

Each project includes strict QC checkpoints, traceable documentation, and publication-quality figures. Our data deliverables are formatted for immediate inclusion in manuscripts or grant submissions.

5. Global Client Trust

Trusted by leading universities, biotech firms, and pharmaceutical R&D teams, CD Genomics continues to serve as a preferred CRO partner for genome structure and transcriptional regulation studies.

Ready to accelerate your research?

Contact CD Genomics to discuss your project or request a customized G-loop Analysis consultation today.

Sample Requirements

Sample Type	Sample Requirements	Notes
Cells	≥ 2 × 10⁵ viable cells (fresh or cryopreserved in cryoprotectant solution)	Frozen cell pellets or cells stored without cryoprotectant are not accepted.
Animal Tissues	≥ 0.5 g internal organs; ≥ 1 g muscle tissue	Frozen animal tissues and tissues preserved in cryoprotectant are acceptable.
Plant Tissues	≥ 0.5 g fresh leaf or stem tissue (Arabidopsis or common crop species)	Frozen plant tissues and actively growing tissues are acceptable.
Species Requirement	Samples must be from species with a reference genome available	Required for accurate sequence alignment and hotspot annotation.

Tip: Avoid repeated freeze–thaw cycles and ensure sterile collection conditions to maintain chromatin integrity and native G4/R-loop structures.

Demo Results Display

Genome browser-style plot showing overlapping G4, R-loop, and G-loop enrichment peaks.

Chromosome map and feature distribution chart showing G-loop signal density across genomic regions.

Volcano plot and motif logo summarizing differential G-loop regions and enriched sequence motifs.

GO/KEGG enrichment bubble chart highlighting biological processes linked to G-loop activity.

Heatmap illustrating correlation and clustering of G-loop sequencing replicates.

1. Peak Detection and Enrichment Profiles

Genome browser snapshots and enrichment plots display the distribution of G4, R-loop, and G-loop signals. Overlapping peaks (purple for G4, teal for R-loop, and green for G-loop) indicate true co-localization regions with high structural confidence.

2. Chromosome and Genomic Feature Distribution

Summary charts show how G-loop signals are distributed across chromosomes and genomic elements such as promoters, enhancers, and gene bodies, providing insight into regulatory hotspot density.

3. Differential Peak and Motif Analysis

Volcano plots highlight significant differential G-loop regions between experimental groups, while motif logos reveal sequence patterns underlying G4 and R-loop co-occurrence.

4. Functional Enrichment (GO/KEGG)

Bubble plots or bar charts illustrate enriched biological processes and pathways, helping link G-loop activity to transcriptional control, DNA repair, or genome maintenance.

5. Correlation and Clustering

Heatmaps and correlation matrices visualize data consistency across replicates, confirming high experimental reproducibility and signal integrity.

FAQs

- What is G-loop analysis and why is it important?
  - G-loop analysis identifies genomic regions where G-quadruplexes (G4s) and R-loops overlap, forming three-stranded nucleic acid structures known as G-loops. These regions play a vital role in transcription regulation, replication stress response, and genome stability. Understanding their distribution helps researchers reveal how RNA–DNA interactions shape gene expression and chromatin architecture.
- How does G-loop Analysis differ from ChIP-seq or DRIP-seq?
  - Unlike ChIP-seq or DRIP-seq, G-loop Analysis uses Cut&Tag chemistry, which performs in situ DNA tagging without crosslinking or sonication. This preserves native G4 and R-loop conformations and produces data with higher resolution, lower background noise, and improved reproducibility. It also requires far fewer cells than traditional immunoprecipitation-based methods.
- What kind of samples are suitable for G-loop Analysis?
  - G-loop Analysis accepts a wide range of sample types, including cultured cells, animal tissues, and plant tissues. The key requirement is that the samples are viable, properly preserved, and from species with available reference genomes. Maintaining chromatin integrity is essential for reliable detection of G4 and R-loop structures.
- What information can researchers obtain from G-loop sequencing data?
  - Researchers receive genome-wide maps of G4 and R-loop signals and integrated G-loop hotspot regions, along with associated motif analyses, GO and KEGG enrichment, and annotated gene lists. These results reveal how G4 and R-loop interactions influence transcriptional regulation, DNA repair pathways, and genome structure.
- Is G-loop Analysis applicable to disease or drug research?
  - Yes. G-loop mapping supports the investigation of diseases involving transcriptional dysregulation or genome instability, such as cancer and neurodegenerative disorders. It also provides valuable insights for drug discovery, particularly for developing small molecules targeting G-quadruplex structures or molecular glue degraders that recognize G-loop motifs.
- Can G-loop Analysis be integrated with other omics data?
  - Absolutely. Results from G-loop Analysis can be integrated with RNA-seq, ATAC-seq, or ChIP-seq datasets to correlate structural features with gene expression, chromatin accessibility, and histone modifications, providing a multi-dimensional view of genome function.
- What support does CD Genomics provide after data delivery?
  - Every project includes detailed bioinformatics reports, visual figures, and expert guidance for data interpretation. Clients receive publication-ready outputs and have the option to discuss results with our scientific team to align findings with their experimental goals.

References:

Bhatt U, Evans CW, Cucchiarini A, Gros J, Iyer KS, Mergny JL, Smith NM. G-quadruplex structural motifs modulate protein-RNA interactions within the transcriptome. Genome Biol. 2025
Lyu J, Shao R, Kwong Yung PY, Elsässer SJ. Genome-wide mapping of G-quadruplex structures with CUT&Tag. Nucleic Acids Res. 2022 Feb 22;50(3):e13. doi: 10.1093/nar/gkab1073. PMID: 34792172; PMCID: PMC8860588.
Maurizio, I., Ruggiero, E., Zanin, I. et al. CUT&Tag reveals unconventional G-quadruplex landscape in Mycobacterium tuberculosis in response to oxidative stress. Nat Commun 16, 7253 (2025).

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