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Enhanced Reduced Representation Bisulfite Sequencing for Assessment of DNA Methylation at Base Pair Resolution
Published on: February 24, 2015
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Low Input Whole-Genome Bisulfite Sequencing Using a Post-Bisulfite Adapter Tagging Approach
Julian R Peat1, Sébastien A Smallwood2,3
1Epigenetics Programme, Babraham Institute, Babraham Research Campus, Cambridge, CB22 3AT, UK.
Methods in Molecular Biology (Clifton, N.J.)
|December 11, 2017
Summary
We developed a new whole-genome bisulfite sequencing method for low cell inputs. This technique enables comprehensive DNA methylation mapping from limited samples, advancing epigenetics research.
Area of Science:
- Epigenetics and Genomics
- Molecular Biology
Background:
- The epigenetic mark 5-methylcytosine regulates gene expression and cell fate.
- High-throughput sequencing allows genome-wide DNA methylation mapping.
- Analyzing limited cell material for DNA methylation remains a challenge.
Purpose of the Study:
- To optimize whole-genome bisulfite sequencing (WGBS) for low cell inputs.
- To enable comprehensive DNA methylation landscape analysis from 100-5000 somatic cells.
Main Methods:
- Developed a post-bisulfite adapter tagging (PSAT) approach for WGBS.
- Bisulfite treatment precedes library generation, fragmenting DNA.
- Adapter ligation via complementary strand synthesis and PCR amplification.
Main Results:
- Successfully performed WGBS on low cell inputs (100-5000 cells).
- Optimized protocol provides comprehensive genome-wide DNA methylation maps.
- Demonstrated feasibility of epigenomic analysis on limited biological material.
Conclusions:
- The PSAT-based WGBS protocol is effective for low-input samples.
- This method overcomes limitations in analyzing sparse cellular material.
- Facilitates epigenomic studies in cell fate transitions and rare cell populations.
