Hypermethylated RASSF1A in maternal plasma:: A universal fetal DNA marker that improves the reliability of noninvasive prenatal diagnosis
Hypermethylated RASSF1A in maternal plasma:: A universal fetal DNA marker that improves the reliability of noninvasive prenatal diagnosis
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DOI：
10.1373/clinchem.2006.074997
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发表时间：
2006-12-01
影响因子：
9.3
通讯作者：
Lo, Y. M. Dennis
中科院分区：
文献类型：
--
作者：
Chan, K. C. Allen;Ding, Chunming;Lo, Y. M. Dennis
背景资料：我们最近发现RASSF1A基因的启动子在胎盘中是高甲基化的，而在母体血细胞中是低甲基化的。这种甲基化模式允许使用甲基化敏感的限制性内切酶消化检测胎盘来源的高甲基化RASSF1A序列在maternal plasma.Methods：我们进行甲基化敏感的限制性内切酶消化后的实时PCR检测胎盘来源的RASSF1A序列在血浆中的28个第一孕期和43个第三孕期孕妇。对54例妊娠早期RhD阴性孕妇进行产前胎儿RhD血型分型，并以RASSF1A高甲基化作为阳性对照进行胎儿DNA检测。酶消化后的血浆RASSF1A基因型与每种情况下的胎儿基因型相同，从而证实其胎儿来源。在54名接受产前胎儿RhD基因分型的孕妇中，有19名在其血浆DNA样本中检测不到RHD序列。胎儿DNA对照RASSF1A在19名妇女中的4名中检测不到。随后的绒毛膜绒毛样本分析显示，这4名妇女与负RHD和RASSF1A信号实际上是携带RhD阳性fetal.Conclusions：高甲基化RASSF1A是一个通用的标记胎儿DNA，是很容易检测到在母体血浆。当应用于产前RhD基因分型时，该标记物允许检测由母体血浆中低胎儿DNA浓度引起的假阴性结果。这种新的标记物也可以应用于许多其他产前诊断和监测方案。(c)2006年美国临床化学协会。
Background: We recently demonstrated that the promoter of the RASSF1A gene is hypermethylated in the placenta and hypomethylated in maternal blood cells. This methylation pattern allows the use of methylation-sensitive restriction enzyme digestion for detecting the placental-derived hypermethylated RASSF1A sequences in maternal plasma.Methods: We performed real-time PCR after methylation-sensitive restriction enzyme digestion to detect placental-derived RASSF1A sequences in the plasma of 28 1st-trimester and 43 3rd-trimester pregnant women. We used maternal plasma to perform prenatal fetal rhesus D (RhD) blood group typing for 54 early-gestation RhD-negative women, with hypermethylated RASSF1A as the positive control for fetal DNA detection.Results: Hypermethylated RASSF1A sequences were detectable in the plasma of all 71 pregnant women. The genotype of plasma RASSF1A after enzyme digestion was identical to the fetal genotype in each case, thus confirming its fetal origin. Nineteen of the 54 pregnant women undergoing prenatal fetal RhD genotyping showed undetectable RHD sequences in their plasma DNA samples. The fetal DNA control, RASSF1A, was not detectable in 4 of the 19 women. Subsequent chorionic villus sample analysis revealed that 2 of these 4 women with negative RHD and RASSF1A signals were in fact carrying RhD-positive fetuses.Conclusions: Hypermethylated RASSF1A is a universal marker for fetal DNA and is readily detectable in maternal plasma. When applied to prenatal RhD genotyping, this marker allows the detection of false-negative results caused by low fetal DNA concentrations in maternal plasma. This new marker can also be applied to many other prenatal diagnostic and monitoring scenarios. (c) 2006 American Association for Clinical Chemistry.