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Rapid screening of the heterogeneity of DNA methylation by single‐strand conformation polymorphism and CE ‐ LIF in the presence of electro‐osmotic flow
Author(s) -
Yu MengHsuan,
Huang YaChi,
Chang PoLing
Publication year - 2014
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.201300502
Subject(s) - dna methylation , methylation , single strand conformation polymorphism , dna , microbiology and biotechnology , epigenetics , bisulfite , bisulfite sequencing , chemistry , biology , polymerase chain reaction , biochemistry , gene , gene expression
DNA methylation is a complex event in epigenetic studies because of both the large C p G islands present upstream of the promoter region and the different distribution of DNA methylation despite similar methylation levels. For this reason, we proposed a fast, cost‐effective method for the screening of DNA methylation based on SSCP and CE ‐ LIF . In this study, the PCR products that were amplified from bisulfite‐treated genomic DNA were denatured at 94°C, followed by immediate chilling in ice water to form the ss DNA . The ss DNA were separated by 1.5% poly(ethylene oxide) ( M avg 8 000 000 Da) in the presence of EOF according to the different conformations represented by their unique methylation states. This result demonstrated that four hepatocellular carcinoma cell lines represented a different heterogeneity of DNA methylation and could be distinguished by SSCP ‐ CE . The results obtained from SSCP ‐ CE also corresponded with those obtained from combined bisulfide restriction analysis and methylation‐sensitive high‐resolution melting analysis. Therefore, the proposed SSCP ‐ CE method may potentially be used for rapid screening for determination of the heterogeneity of DNA methylation in further epigenetic studies and clinical diagnosis.