RASSF1A在无损性孕妇外周血浆中胎儿ABO基因分型中的运用

Application of RASSF1A to Noninvasive Fetal ABO Genotyping in Pregrnant Peripheral Plasma

目的:探讨甲基化表观遗传标志RASSF1A在O型孕妇外周血浆中胎儿ABO基因分型中的运用,以无损性预测产前胎儿ABO血型。方法:首先提取O型孕妇外周血浆游离DNA,扩增SRY及甲基化RASSF1A作为胎儿DNA存在的标志;检测血浆游离DNA中B及非O预测胎儿ABO血型;并将基因分型结果与胎儿分娩后血清学结果进行比对,以评价该方法的准确性。结果:20例标本中,11例检测出SRY,检测结果与胎儿出生后性别相符;其余9例标本酶切后均扩增出RASSF1A,证实胎儿游离DNA存在。对血浆游离DNA进行ABO基因检测,20例标本中8例标本同时扩增出非O和B,提示为B血型;5例标本只扩增出非O,未扩增出B,提示为A型;7例标本均未扩增出非O及B,提示为O型。上述预测结果与胎儿出生后ABO血清学血型一致。结论:初步建立了基于RASSF1A的O型孕妇血浆中游离DNA中胎儿ABO基因的检测程序,可用于临床无损性产前胎儿ABO血型的预测。

Objective:To investigate the feasibility of noninvasive fetal ABO genotyping based on RASSF1 A gene with circulating cell-free fetal DNA(cffDNA) from maternal plasma.Methods:DNA was extracted from the 0 group pregnant plasma,and the presence of cffDNA was confirmed by fetal DNA maker SRY and RASSF1 A.B and non-0 were detected by real-time PCR,and the genotyping results were evaluated by using the serologic tests for ABO phenotyping.Results:Among the samples of 20 cases,the SRY was found in 11 cases by detecteion,the detection results were consistent with sex of infants after delivery;the RASSF1 A was amplified all in samples of other 9 cases after BstU1 cleavage,which confirmed existance of cffDNA.The ABO gene detection of cffDNA in plasma showed that out of 20 samples,both non O and B were amplified simultancously in 8 cases,suggesting the B blood group;the non O was amplified,but the B was not amplified only in 5 cases,suggesting A blood group,the non O and B both were not amplified in samples of 7 cases,suggesting 0 blood group.The above-mentioned detection results were consistent with new born ABO blood group by serological test.Conclusion:The proposed protocol for the detection of fetal ABO based on RASSF1 A gene by using fetal DNA from maternal plasma can be used for noninvasive prenatal diagnosis of fetal ABO blood group.