An Assay for the Activity of DNA Methylase Based on a Hairpin Fluorescence Molecular Probe and Its Application in Drug Selection

Abstract

A new activity assay for DNA methylase based on a hairpin molecular probe was proposed in this paper. The recognition site of DNA methylase and corresponding restriction endonuclease was designed in the stem part of the probe, tetramethylrhodamine (TAMRA) was attached at the 5' terminus and its fluorescence was quenched by 4-(4'-dimethylaminophenylazo)benzoic acid (DABCYL) linked at the 3' terminus. The unmethylated probe can be cut in the recognition site by restriction endonuclease, causing the restoration of the fluorescence of TAMRA. Therefore, the activity of methylase would be analyzed according to the degree of fluorescence restoration. Based on this assay, the influence of anti-tumor drugs on the activity of methylase was investigated by adding drugs in the methylation reaction. This method can provide a potential in selecting proper drugs for tumors caused by altered gene methylation pattern.

Key words: molecular probe, DNA methylase, fluorescence, drug selection

Cite this article

ZHOU Xing-Wang^1,2; LI Jun^1,2,3; WANG Ke-Min*^,1,2,3; TAN Wei-Hong^1,2,3;YANG Xiao-Hai^1,2; MENG Xiang-Xian^2,3; YAN Hong-Fei^{. An Assay for the Activity of DNA Methylase Based on a Hairpin Fluorescence Molecular Probe and Its Application in Drug Selection[J]. Acta Chimica Sinica, 2006, 64(20): 2096-2100.}

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发夹型荧光分子探针用于DNA甲基化酶的活性分析及其在药物筛选中的应用