Citation: MU Xiaoli, ZHANG Jie, PENG Siyuan, WANG Xiaoxue, SHEN Heqing. Simultaneous analysis of global DNA methylation and hydroxymethylation in tissues by liquid chromatography-tandem mass spectrometry[J]. Chinese Journal of Chromatography, ;2014, 32(7): 682-686. doi: 10.3724/SP.J.1123.2014.03025 shu

Simultaneous analysis of global DNA methylation and hydroxymethylation in tissues by liquid chromatography-tandem mass spectrometry

1.
Key Laboratory of Urban Environment and Health, Institute of Urban Environment, Chinese Academy of Sciences, Xiamen 361021, China

Corresponding author: ZHANG Jie,
Received Date: 18 March 2014
Available Online: 15 May 2014
Fund Project: 国家自然科学基金项目(21177123) (21177123)CAS/SAFEA创新研究国际合作项目(KZCX2-YW-T08). (KZCX2-YW-T08)

Measuring global DNA methylation and hydroxymethylation is important in researches of effect and mechanism of environmental pollutants exposure. A method based on liquid chromatography-electrospray ionization tandem mass spectrometry (LC-MS/MS) was developed to simultaneously determine global DNA methylation and hydroxymethylation level in biological tissues. DNA was extracted from tissues and converted into single nucleotide via enzyme digestion. Liquid chromatography coupled to tandem mass spectrometry was used to measure the concentrations of 5-methylcytidine, 5-hydroxymethylcytosine and deoxyguanosine, which were used to calculate global DNA methylation ratios and hydroxymethylation ratios. The results showed that the correlation coefficients were higher than 0. 99. The limit of detection (LOD, S/N=3) and the limit of quantification (LOQ, S/N=10) of 5-methylcytidine were 0.015 and 0.045 ng/mL, respectively. They reached 0.001 and 0.003 ng/mL for 5-hydroxymethylcytosine, and were 0.2 and 0.6 ng/mL for deoxyguanosine. The developed method was further successfully applied to investigate global DNA methylation and hydroxymethylation alteration in liver and cerebellum of rats exposed to arsenic via drinking water. This approach could quantitatively detect 5-methylcytidine, 5-hydroxymethylcytosine and deoxyguanosine with high sensitivity, repeatability and stability. Our study provided a means to simultaneously analyze global DNA methylation and hydroxymethylation.
- liquid chromatography-tandem mass spectrometry (LC-MS/MS),
- 5-methylcytidine (5mC),
- 5-hydroxymethylcytosine (5hmC),
- tissues
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沈阳化工大学材料科学与工程学院沈阳 110142