GRO-seq
GRO-seq
Global nuclear Run-On sequencing (GRO-seq) is a specialized method designed to measure nascent RNA (Core L J, et al. Science, 2008.), enabling the systematic identification of enhancer RNAs (eRNAs). The protocol involves the rapid cryopreservation of cells to arrest transcriptional complexes within the nucleus. Following nuclei isolation, a "run-on" buffer is introduced to resume RNA transcription in the presence of Br-UTP. The resulting BrU-labeled nascent RNA is immunopurified using anti-BrdU antibodies and subjected to high-throughput sequencing.
Because changes in nascent RNA directly reflect immediate transcriptional regulation mechanisms, GRO-seq technology allows for the observation of genome-wide active transcription. It generates high-resolution maps depicting the position, density, and orientation of active RNA Polymerase II.
Tzerpos P, et al. Methods Mol Biol. 2021.
Workflow
Sample Requirements
Type & Input
Cells，≥1×10⁷ cells/sample
Validated for Human, Mouse, and Rat. (Other species require a feasibility assessment).
Applications
1. Supersedes conventional RNA-seq by enabling precise detection of gene expression during active transcription；
2. Precisely determines transcription start sites (TSSs) and transcriptional orientation；
3. Enables identification of direct transcriptional targets；
4. Facilitates discovery of novel transcripts, including non-coding RNAs；
5. Enables identification of enhancer RNAs (eRNAs)；
Bioinformatics Analysis
1. Data QC: adapter trimming, low-quality read filtering, and contamination removal.
2. Genome alignment and transcriptome-wide read distribution analysis.
3. Meta-analysis of read enrichment at TSSs.
4. Analysis of transcriptional pausing.
5. eRNA identification and analysis.
6. Differential expression analysis.
7. GO/KEGG analysis of differentially expressed genes.
Visualization of GRO-seq data (bed files) in the UCSC Genome Browser
Demo
Histogram of TSS distances
GRO-seq signal enrichment centered on enhancer regions across samples
Boxplot illustrating changes in the Pausing Index
Heatmap of condition-dependent transcriptional changes in active genes surrounding the TSS
Histogram showing transcriptional variation between replicate samples
EPIBIOTEK® Project Publications
Wang WL, Song ZX, Bu SM, et al. Ythdc1-p300-Klf5 Complex-Mediated Golgi Dysfunction Promotes Aortic Aneurysm. Adv Sci (Weinh). 2026;13(4):e12116. doi:10.1002/advs.202512116