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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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Efficient and Sensitive Bisulfite-Free Methylation Detection Approach for Low-Abundance and Highly Fragmented cfDNA.
Linqing Zhen1,2, Hao Yang1,2, Yizhou Huang1,2
1School of Biomedical Engineering, Shanghai Jiao Tong University, Shanghai 200030, P. R. China.
Analytical Chemistry
|September 26, 2025
Summary
A new method called dRE-STEM detects DNA methylation in cell-free DNA (cfDNA) for early cancer diagnosis. This technique improves sensitivity for fragmented cfDNA, outperforming traditional protein markers in colorectal cancer detection.
Area of Science:
- Biochemistry
- Molecular Biology
- Oncology
Background:
- Circulating cell-free DNA (cfDNA) is a promising noninvasive biomarker for liquid biopsy.
- Abnormal DNA methylation in cfDNA is crucial for cancer screening and early diagnosis.
- Current methylation detection methods struggle with low cfDNA abundance and fragmentation.
Purpose of the Study:
- To develop a highly sensitive cfDNA methylation detection method.
- To establish a bisulfite conversion-free approach for fragmented cfDNA.
- To validate the diagnostic value of the novel method for colorectal cancer (CRC).
Main Methods:
- Combined double restriction enzyme digestion with specific terminal-mediated polymerase chain reaction (STEM-PCR).
- Developed the dRE-STEM method for cfDNA methylation detection.
- Conducted a case-control study to assess diagnostic accuracy for CRC.
Main Results:
- The dRE-STEM method detected methylation ratios as low as 3% with 4 ng cfDNA input.
- Achieved 80.2% sensitivity and 80.9% specificity for CRC detection.
- Significantly outperformed conventional protein markers in diagnostic accuracy.
Conclusions:
- The dRE-STEM method enables accurate methylation quantification in low-abundance, fragmented cfDNA.
- This bisulfite conversion-free approach is suitable for routine clinical practice.
- Offers a promising tool for noninvasive cancer screening and early diagnosis.
