Related Experiment Video
Updated: Jul 8, 2026
08:08
Formaldehyde-assisted Isolation of Regulatory Elements to Measure Chromatin Accessibility in Mammalian Cells
Published on: April 2, 2018
Quantitative differences in chromatin accessibility across regulatory regions can be directly compared in distinct
Mark Cruickshank1, Emily Fenwick, Lawrence J Abraham
1Biochemistry and Molecular Biology, School of Biomedical, Biomolecular and Chemical Sciences, The University of Western Australia, 35 Stirling Highway, Crawley, 6009 WA, Australia.
Biochemical and Biophysical Research Communications
|January 1, 2008
Summary
Chromatin accessibility correlates with human CR2 gene expression. The CHART-PCR assay revealed distinct accessibility patterns in expressing B-cells compared to non-expressing cells, validating its use for comparing cell types.
Area of Science:
- Molecular Biology
- Epigenetics
- Gene Regulation
Background:
- Transcriptional activation in eukaryotes involves chromatin structure alterations.
- Chromatin accessibility at regulatory sites can vary while other genomic regions remain unchanged.
Purpose of the Study:
- To examine the correlation between CR2 gene expression and chromatin accessibility in various human cell lines.
- To validate the CHART-PCR assay for comparing chromatin accessibility across different cell types.
Main Methods:
- Utilized the Chromatin Accessibility Real-Time PCR (CHART-PCR) assay.
- Employed micrococcal nuclease (MNase) as an accessibility agent.
- Analyzed chromatin accessibility of the human CR2 gene promoter in U937, REH, Ramos, and Raji cell lines.
Main Results:
- CHART-PCR demonstrated site-specific chromatin accessibility, consistent across cell lines for GAPDH and beta-actin promoters.
- Developmentally restricted genes PAX-7 and SP-A2 promoters showed consistently reduced chromatin accessibility.
- Significantly enhanced chromatin accessibility was observed in the CR2 promoter regions (-289 to -101 and -115 to -12) of CR2-expressing B-cells (Ramos, Raji) compared to non-expressing cells (U937, REH).
Conclusions:
- CHART-PCR assays effectively detect correlations between chromatin accessibility and gene expression.
- Chromatin accessibility is specifically altered at the CR2 promoter in expressing B-cells.
- The study validates the utility of CHART-PCR for comparative analyses of chromatin accessibility across different cell types.
Related Concept Videos
02:36Chromatin Immunoprecipitation- ChIP
Chromatin immunoprecipitation, or ChIP, is an antibody-based technique used to identify sites on DNA that bind to transcription factors of interest or histone proteins. It also helps determine the type of histone modifications such as acetylation, phosphorylation, or methylation.
Types of ChIP
ChIP can be divided into two types - X-ChIP and N-ChIP. X-ChIP involves in vivo cross-linking of histones and regulatory proteins to DNA, fragmenting the DNA by sonication, and isolating the protein-DNA...
Types of ChIP
ChIP can be divided into two types - X-ChIP and N-ChIP. X-ChIP involves in vivo cross-linking of histones and regulatory proteins to DNA, fragmenting the DNA by sonication, and isolating the protein-DNA...
02:35Chromatin Position Affects Gene Expression
Chromatin is the massive complex of DNA and proteins packaged inside the nucleus. The complexity of chromatin folding and how it is packaged inside the nucleus greatly influences access to genetic information. Generally, the nucleus' periphery is considered transcriptionally repressive, while the cell's interior is considered a transcriptionally active area.
Topologically Associated Domains (TADs)
The 3-dimensional positioning of chromatin in the nucleus influences the timing and level of...
Topologically Associated Domains (TADs)
The 3-dimensional positioning of chromatin in the nucleus influences the timing and level of...
01:32Chromatin Modification in iPS Cells
Chromatin modification alters gene expression; therefore, scientists can add histone-modifying enzymes, histone variants, and chromatin remodeling complexes to somatic cells to aid reprogramming into pluripotent stem (iPS) cells.
Compact chromatin makes reprogramming difficult. Enzymes, such as histone demethylases and acetyltransferases, are often added during reprogramming to loosen the chromatin, making the DNA more accessible to transcription factors. Molecules that inhibit histone...
Compact chromatin makes reprogramming difficult. Enzymes, such as histone demethylases and acetyltransferases, are often added during reprogramming to loosen the chromatin, making the DNA more accessible to transcription factors. Molecules that inhibit histone...
01:01Euchromatin
The extent of chromatin compaction can be studied by staining chromatin using specific DNA binding dyes. Under the microscope, the dense-compacted regions take up more dye, appearing darker, while the less-compact areas take up less dye and appear lighter. Based on the compaction level, chromatins are classified into two primary forms – euchromatin and heterochromatin.
Euchromatin is the less dense region of the chromatin and stains lighter. Euchromatin contains histone H3 extensively...
Euchromatin is the less dense region of the chromatin and stains lighter. Euchromatin contains histone H3 extensively...
02:38Heterochromatin
The extent of chromatin compaction can be studied by staining chromatin using specific DNA binding dyes. Under the microscope, the dense-compacted regions that take up more dye are called heterochromatin. Heterochromatin is further classified into two forms – constitutive heterochromatin and facultative heterochromatin.
Constitutive heterochromatin: It is a highly compact region of chromatin that is mostly concentrated in the centromere and telomere. Unlike euchromatin, the amino acid at 9th...
Constitutive heterochromatin: It is a highly compact region of chromatin that is mostly concentrated in the centromere and telomere. Unlike euchromatin, the amino acid at 9th...