High-throughput assay of DNA methylation based on methylation-specific primer and SAGE.
Mapping of genomic DNA methylation is a dispensable part of functional genome. We have developed a novel method based on methylation-specific primer and serial analysis of gene expression, called MSP-SAGE, with potential of high-throughput quantification of genomic DNA methylation. We used a 6-mer methylation-specific primer to extend the methylated CpG sequences other than non-methylated CpG sequences. The 17 bp tags contained methylated CpG sequence, which were obtained from extended methylation sequence by digestion of restriction endonuclease, and then the tags were concatenated and cloned for sequencing. We can identify the locations of methylation according to the sequences of tags and quantify the methylation status from the frequency of the tags. MSP-SAGE has a good linearity in a broad methylation range from 5% to 100% with good accuracy and high precision. The proof-of-principle study shows that MSP-SAGE is a reliable high-throughput assay for quantification of DNA methylation.
Wang
Xianliang
X
MOE Key Lab of Environment and Health, School of Public Health, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
Zhang
Chi
C
Zhang
Lijun
L
Wang
Xiaoli
X
Xu
Shunqing
S
D000483:Alleles; D018899:CpG Islands / Q000235:genetics; D019175:DNA Methylation; D017931:DNA Primers / Q000235:genetics*; D020869:Gene Expression Profiling
D017931:DNA Primers