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Single-molecule DNA methylation detection with fluorescent AdoMet analogues

DNA methylation plays a crucial role in gene expression regulation, and aberrant methylation patterns are implicated in cancer biogenesis. Current single-molecule methods focus primarily on hypermethylation detection; however, many oncogenes are hypomethylated during cancer development. For this reason, Gelboa et al. developed a method for labeling and single-molecule quantification of multiple unmethylated cytosine-guanine dinucleotides (CpGs).

The authors took advantage of DNA methyltransferases (MTases) that catalyze the transfer of a methyl group from S-adenosyl-L-methionine (AdoMet) to cytosine or adenine at specific DNA sequences. Fluorescent analogues of AdoMet were made, using TAMRA or CF™640R and these fluorescent reporters were then used for MTase-catalyzed labeling of unmethylated CpG sites on DNA molecules. An ultrasensitive electro-optical nanopore detection system was employed to produce highly quantitative single-molecule fluorescence measurements during the passage of each DNA molecule through a nanopore.

To read the original article, click here.

T. Gilboa, C. Torfstein, M. Juhasz, A. Grunwald, Y. Ebenstein, E. Weinhold, and A. Meller. Single-Molecule DNA Methylation Quantification Using Electro-optical Sensing in Solid-State Nanopores. ACS Nano 2016, 10, 8861−8870. DOI: 10.1021/acsnano.6b04748.

CF640R phalloidin

 

Biotium offers bright and photostable fluorescent CF™ dyes with a full selection of chemically reactive functional groups for labeling biomolecules. Visit the CF™ Dye technology page to learn more.