Circular Chromosome Conformation Capture Sequencing (4C-Seq ) in Primary Adherent Cells
Judith Marsman1, Robert C Day2, Gregory Gimenez3
1Department of Cardiology, Division Heart & Lungs, University Medical Centre Utrecht, Utrecht, The Netherlands. j.marsman@umcutrecht.nl.
Methods in Molecular Biology (Clifton, N.J.)
|February 1, 2022
Summary
We optimized circular chromosome conformation capture sequencing (4C-seq) for primary adherent cells to map genome-wide chromatin interactions. This improved protocol enhances gene regulation studies by efficiently capturing 3D genome structures.
Area of Science:
- Genomics
- Molecular Biology
- Epigenetics
Background:
- The three-dimensional genome structure is crucial for gene regulation.
- Chromosome conformation capture (3C) techniques identify physical interactions between genomic loci.
- Circular chromosome conformation capture sequencing (4C-seq) maps interactions from a single viewpoint.
Purpose of the Study:
- To describe an optimized 4C-seq protocol for primary adherent cells.
- To improve the efficiency of the initial digestion step in 4C-seq.
- To provide a comprehensive protocol applicable to various cell and tissue types.
Main Methods:
- Optimization of the 4C-seq protocol for primary adherent cells.
- Utilizing a standard DNA library preparation method with a commercial kit.
- Detailed description of data processing steps for 4C-seq analysis.
Main Results:
- The optimized protocol shows improved efficiency in the critical first digestion step for adherent cells.
- The protocol is effective for identifying chromatin interactions in a "one versus all" manner.
- Successful application to primary adherent cells and potential applicability to other cell types.
Conclusions:
- The described 4C-seq protocol is a valuable tool for studying 3D genome organization.
- This method facilitates the investigation of gene regulation through chromatin interactions.
- The protocol provides a robust approach for mapping genome architecture in diverse biological samples.
