Determination of global DNA methylation in human leukocyte by isocratic cation exchange high-porformance liquid chromatography

VernacularTitle:离子交换色谱法检测人血白细胞基因组DNA整体甲基化水平
Author: Zheng LI ; Ying LI ; Yongjia YANG ; Fuyou LIU ; Wuqiu LI
Publication Type:Journal Article
Keywords: Chromatography,ion exchange; Genome; DNA methylation
From: Journal of Chinese Physician 2010;12(1):55-58
CountryChina
Language:Chinese
Abstract: Objective Improving the existed HPLC methods in order to detect the levels of global DNA methylation rapidly, stably and conveniently. Methods The HP1100 high performance liquid chrom-atographic (HPLC) system was used in this study. The analytic column was Macherey-Nagel (MN) EC 250/4.6 Nucleosil 100-5SA (250mm x 4. 6mm, 5u,m) cation exchange chromatographic column. We used 60mM acetic acid + 15% acetonitrile as mobile phase (adjust to pH =4. 6 by NaOH). The flow rate was 1. 0 ml/min, detective UV wavelength was set to 276 nm and column temperature was set to 28℃. The injection volume was 50μl. The global DNA methylation was expressed as 5mdC/(dC +5mdC) x 100%. Results Under these conditions, we can isolate dC and 5mdC completely in ten minutes. The level of leukocyte global DNA methylation in healthy people is (4. 389 ±0. 0159) %. Conclusions This method can determine the levels of global DNA methylation rapidly, and it can be widely applied in some laboratories.