孕妇血浆中胎源性RASSF1A基因在无损性产前诊断中的应用研究

Detection of Fetal RASSF1A Gene in Maternal Plasma for Noninvasive Prenatal Diagnosis

本研究旨在探讨运用非性别依赖的甲基化表观遗传学标记RASSF1A基因作为孕妇血浆中循环游离胎儿DNA存在的通用性标志的可行性。采用2种方法提取孕妇血浆游离DNA,选用其中效果较好的用磁珠法提取20例标本的游离DNA;采用RT-PCR方法扩增SRY基因以及甲基化酶处理后的RASSF1A基因,且对酶处理后RASSF1A基因的扩增体系进行条件优化。结果表明,在20例标本中11例检测有SRY基因,且与胎儿出生后性别相符;选用优化后的PCR体系扩增甲基化酶处理后的RASSF1A基因,在18例标本中检测有RASSF1A基因,在2例标本中未检测到此基因。结论:甲基化RASSF1A基因与SRY基因相比,不受胎儿性别限制,可作为广泛性胎儿特异性表观遗传学标记物,用于临床无损性产前诊断。

The aim of this study was to investigate the feasibility of using RASSF1A gene as a universal fetal marker in maternal plasma. Two methods of circulating cell-free fetal DNA （cffDNA） extacted from maternal plasma were compared. The better one was chosen for extraction of cffDNA in the 20 pregnant samples. The SRY gene and the RASSF1A gene treated with methylation-sensitive restriction enzyme were amplificated by RT-PCR and the PCR system was optimized. The results showed that the SRY gene was found in 11 out of the 20 pregnant samples, which was consistent with the postnatal sex. Using the optimized PCR system, the specifically amplified fetal-associated methylated RASSF1A gene was found after trentment with BstUI in 18 of the 20 pregnant samples, while the 2 samples failed in detection. It is concluded that the methylated fetal-specific RASSF1A gene can be used as a univeisal fetal marker for the presence of cffDNA in maternal plasma without fetal gender restrictions. So, it can be used for noninvasive prenatal diagnosis.