甲基化特异性多重连接依赖探针扩增技术在胎盘甲基化研究中的应用

Application of methylation-specific multiplex ligation-dependent probe amplification for the study of DNA methylation in placenta tissues

目的探讨甲基化特异性多重连接依赖探针扩增技术（methylation—specific multiplex ligation-dependent probe amplification，MS-MLPA）在胎盘甲基化研究中的可行性。方法应用MS-MI。PA及亚硫酸氢盐测序两种方法对9个正常整倍体胎盘样本及13名非孕妇女外周血进行甲基化检测，并计算候选的CG1149、HLCS、CG1113、ACTB4个基因的甲基化率。结果在13名非孕妇女的外周血标本中，当消化内参完全消化时，ACTB基因的甲基化率波动于0～0．1之间，0．1为MS-MLPA检测甲基化率的截断值；在9个胎盘组织样本中，MS-MLPA与亚硫酸氢盐测序方法所测得的HLCS、CGH13、CG1149和ACTB4个基因的甲基化率结果一致。结论MS-MLPA检测甲基化率结果可靠，可代替亚硫酸氢盐测序用于胎盘的甲基化研究。

Objective To study the feasibility of using methylation-specific multiplex ligationdependent probe amplification （MS-MLPA） for the detection of DNA methylation in placenta tissue. Methods For blood cells from 13 non-pregnant women and 9 euploid placenta, the ratios of DNA methylation were evaluated for 4 genes including CGI149, CGIl13, HLCS and ACTB with MS-MLPA and bisulfite sequencing, respectively. Results The methylation ratio of the ACTB gene was 0-0. 1 for the blood cells when the digestion control was completely digested. The cutoff value for the methylation ratio of MS- MLPA has been determined as 0.1. For the 9 placenta samples, results of MS-MLPA and bisulfite sequencing were concordant for all of the four genes. Conclusion MS-MLPA is an effective alternative to bisulfite sequencing for the assessment of methylation ratios in placental tissues.