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The Human CUT&Tag Assay Kit for Illumina is a next-generation chromatin mapping technology that has transformed how laboratories explore DNA-protein interactions. With this kit, researchers can achieve single-base resolution of chromatin-bound targets using low cell numbers and streamlined library preparation optimized for Illumina® sequencing platforms.

As epigenomic mapping becomes increasingly important in molecular biology, CUT&Tag (Cleavage Under Targets and Tagmentation) has become the go-to method for high-resolution chromatin profiling with reduced background noise. Unlike conventional ChIP-seq protocols, CUT&Tag requires significantly less sample input and avoids formaldehyde crosslinking and sonication.

Understanding CUT&Tag Technology

The CUT&Tag technique is based on tethering a protein A/G-fused Tn5 transposase directly to chromatin-bound proteins via specific antibodies. Upon activation, the enzyme cuts DNA at the protein binding sites and integrates sequencing adapters, enabling direct library generation.

This targeted approach drastically reduces non-specific signal, a key advantage noted by research groups at Harvard University and Stanford School of Medicine. The protocol is now routinely used in labs studying histone modifications, transcription factors, and nucleosome positioning.

Key Features of the Human CUT&Tag Assay Kit for Illumina

• ✅ Ultra-low input (as few as 5,000 cells)

• ✅ Direct adapter insertion using Tn5 tagmentation

• ✅ Antibody-guided specificity

• ✅ Complete compatibility with Illumina NextSeq and NovaSeq

• ✅ Streamlined magnetic bead purification

• ✅ Library prep in under 8 hours

Protocol Summary

1. Cell Permeabilization – Maintains native chromatin structure.

2. Antibody Binding – Target-specific antibody binds to chromatin.

3. pA/G-Tn5 Binding – Fusion enzyme localizes to antibody.

4. Tagmentation – Enzyme cleaves and inserts Illumina adapters.

5. PCR Amplification – Final libraries are ready for NGS.

The University of Washington Epigenetics Program has published successful adaptations of this protocol across multiple human cell lines and tissue samples.

Application Areas of the Human CUT&Tag Assay Kit

Histone Modification Analysis

One of the most common uses of CUT&Tag is profiling histone post-translational modifications. Antibodies against H3K27ac, H3K4me3, or H3K27me3 can pinpoint enhancer regions and repressed chromatin, crucial for studying regulatory elements in the human genome.

Researchers at Cold Spring Harbor Laboratory have successfully used this technique to map enhancer landscapes in neuronal differentiation models.

Transcription Factor Binding

Unlike ATAC-seq, which measures accessibility, CUT&Tag directly visualizes where transcription factors bind DNA. This is vital for studies involving developmental gene networks, signaling pathways, and cell fate decisions.

The Jackson Laboratory integrates CUT&Tag with CRISPR screening and RNA-seq to dissect gene regulation across different human cell types.

Chromatin Remodeling

CUT&Tag is increasingly utilized to analyze chromatin remodelers such as BRG1, CHD4, or HDAC complexes. The NIH 4D Nucleome project encourages CUT&Tag applications for understanding nuclear architecture dynamics.

Performance Benchmarks

CUT&Tag data quality is significantly higher than that of ChIP-seq in terms of:

These metrics are routinely validated by academic cores such as the University of Michigan Epigenomics Core and the Broad Institute.

Integration with Illumina Platforms

This kit is fully optimized for Illumina platforms including:

• NextSeq 2000 (Illumina NextSeq)

• NovaSeq 6000 (Illumina NovaSeq)

• iSeq and MiniSeq for pilot studies

The preloaded adapter design matches Illumina flow cells, ensuring consistent cluster generation and optimal Q-scores for downstream bioinformatics pipelines.

Data Analysis Compatibility

After sequencing, CUT&Tag data can be processed using open-source software from institutions like:

• ENCODE DCC Pipeline

• Bowtie2 for alignment

• deepTools

• IGV Browser

Advanced researchers may integrate data with RNA-seq, ATAC-seq, or Hi-C datasets for comprehensive regulatory landscape mapping.

Single-Cell and Multi-Omic Applications

Adaptations of the kit allow for single-cell CUT&Tag (scCUT&Tag), making it ideal for resolving chromatin profiles at individual cell resolution. This is especially valuable for identifying cell state transitions and rare subpopulations.

Recent developments from Caltech and UCSF demonstrate how scCUT&Tag can be combined with single-cell RNA-seq to dissect epigenetic control of cell identity.

QC, Reproducibility, and Kit Validation

Each kit lot is validated against positive control antibodies, ensuring consistent signal and library size. Results are benchmarked against ENCODE datasets and metrics published by the NIH Epigenomics Roadmap.

Common QC indicators:

• Library size: 300–700 bp

• Mapping efficiency: >90%

• Duplication rate: <5%

• Peak resolution: ±50 bp

Advantages for Human Research Labs

This kit has been widely adopted in human-focused epigenomic studies including:

• Developmental biology at University of Pennsylvania

• Oncology research at Fred Hutchinson Cancer Center

• Immunology studies at NIH Vaccine Research Center

Its low-input workflow, reproducibility, and Illumina readiness make it a superior choice over ChIP-seq or DNase-seq alternatives.

Kit Contents

Each kit includes a detailed protocol and optimization guide, adapted from UCLA’s Epigenomics Program.

Final Thoughts

The Human CUT&Tag Assay Kit for Illumina is a refined, research-grade solution that makes genome-wide chromatin profiling accessible, scalable, and accurate. Its advantages over traditional approaches, compatibility with Illumina sequencers, and robust scientific validation make it a key resource for labs focusing on gene regulation, transcription dynamics, and epigenetic landscapes.

With ongoing support from platforms like NCBI, PubMed, CDC Genomics, and NIH, this assay format will continue to power human genomic discovery for years to come.