Microarray-based Ms-SNuPE: Near-quantitative analysis for a high-throughput DNA methylation
- 作者：Zhixiang Wu ; Junfeng Luo ; Qinyu Ge ; Zuhong Lu
- 关键词：Methylation ; Single base extension ; Tag ; Microarray ; CpG dinucleotides ; Methylation-specific PCR
- 刊名：Biosensors and Bioelectronics
- 出版年：2008
- 期刊代码：12_09565663
- 类别：ch
- 出版时间：15 April 2008
- 卷：23
- 期：9
- 页码：1333-1339
- 文件大小：546 K
摘要
Aberrant DNA methylation of CpG site in the gene promoter region has been confirmed to be closely associated with carcinogenesis. In the present study, a microarray-based methylation-sensitive single-nucleotide primer extension (Ms-SNuPE) for parallel detecting changes of DNA methylation in cancer was developed. After modification by sodium sulfite, the unmethylated cytosine in the genomic DNA is converted to uracil while leaving the 5-methylcytosine unchanged, which can be detected by bifunctional primer carrying a unique sequence tag in addition to a locus-specific sequence. Because each locus has a distinct tag, the detecting reactions can be performed in a highly multiplexed fashion and the resulting product then be hybridized to the reverse complements of the sequence tags arrayed on a glass slide for methylation analysis. The calibration curves with the correlation coefficient >0.97 were established, which suggested that the method could be used in near-quantitative DNA methylation analysis. Two breast tumor-related genes (E-cad and p16) are successfully analyzed by two group primers (22 primers total), and the results are compatible with that of methylation-specific PCR (MSP). Our research proved that the method is simple and inexpensive, and could be applied as a high-throughput tool to quantitatively determine methylation status of the investigated genes.