用于DNA甲基化高通量分析的甲基化CpG岛扩增与微阵列技术（MCAM）

Methylated CpG Island Amplification and Microarray (MCAM) for High-Throughput Analysis of DNA Methylation.

作者信息

Estécio Marcos R H, Yan Pearlly S, Huang Tim H-M, Issa Jean-Pierre J

机构信息

Department of Leukemia, University of Texas M.D. Anderson Cancer Center, Houston, TX 77030, USA.

出版信息

CSH Protoc. 2008 Mar 1;2008:pdb.prot4974. doi: 10.1101/pdb.prot4974.

DOI:10.1101/pdb.prot4974

PMID:21356790

Abstract

INTRODUCTIONThis protocol describes the use of methylated CpG island amplification (MCA) in combination with a microarray platform to analyze genome-wide DNA methylation in a high-throughput fashion. In this approach, termed MCAM, methylated CpG islands are selectively targeted using oligonucleotide adaptors after two rounds of digestion with a combination of methylation-sensitive and methylation-insensitive nucleases. They are then amplified using PCR. The resulting amplicons, representing the methylated fraction of the genome, are labeled with fluorochromes. Subsequently, a comparative hybridization of reference and test samples (typically normal and tumor DNA specimens) is done on a microarray platform.

摘要

引言

本方案描述了甲基化CpG岛扩增（MCA）与微阵列平台相结合，以高通量方式分析全基因组DNA甲基化的方法。在这种称为MCAM的方法中，先用甲基化敏感和甲基化不敏感的核酸酶组合进行两轮消化，然后使用寡核苷酸接头选择性靶向甲基化的CpG岛。接着通过聚合酶链反应（PCR）进行扩增。得到的扩增子代表基因组的甲基化部分，用荧光染料标记。随后，在微阵列平台上对参考样品和测试样品（通常是正常和肿瘤DNA样本）进行比较杂交。