孕妇外周血浆胎儿DNA的STR检测

Detection of fetal short tandem repeat genotype in maternal plasma by multiplex fluorescent polymerase chain reaction

目的探讨利用孕妇血浆中游离胎儿DNA进行遗传病产前基因诊断的可行性。方法用QIAamp DNA Kit抽提孕妇血浆DNA,应用AmpFl STR profiler试剂盒扩增9个(D3S1358,VWA,FGA,D5S818,D13S317,D7S820,D8S1179,D21S11,D18S51)具有高度多态性的短串联重复序列(short tandem repeat,STR)位点,以多重荧光PCR方法对不同孕期的36份孕妇血浆标本中DNA进行STR等位基因扩增,同时扩增孕妇丈夫外周血DNA。PCR产物经ABI Prism377序列分析仪电泳后,用基因扫描软件进行分析,以胎儿父源性STR等位基因在孕妇血浆中DNA的检出确认胎儿DNA存在。结果孕早期4份(4/6)、孕中期19份(19/20)、孕晚期9份(9/10)检出胎儿父源性等位基因,即胎儿DNA。4份样本未检到胎儿DNA。结论应用多重荧光PCR方法对孕妇血浆中DNA进行STR多态位点的复合扩增,可获得男性和女性胎儿DNA信息,可用于无创伤性产前诊断。

Objective To study the feasibility of using free fetal DNA in maternal plasma for non-invasive prenatal diagnosis.Methods DNA from maternal plasma sample was extracted using QIAamp DNA kit.Using AmpF1 STR profiler box to detect the gen otype of 36 pregnant women at different pregnancy plasma samples by multiplex fluorescent PCR at nine different polymorphic short tandem repeat(STR)loci(D3S1358,VWA,FGA,D5S818,D13S317,D7S820,D8S1179,D21S11,D18S51).Their husband’s DNA isolated from whole blood samples were detected at the same time.The PCR products were electrophoresized by ABI Prism 377 sequencer,the results of electrophoresis were analysed by Genscan.We can confirm the presence of fetal DNA in maternal plasma by that paternally inherited fetal alleles were detected.Results Paternally inherited fetal alleles in 4 cases in early pregnancy(4/6),19 cases in middle pregnancy(19/20)and 9 cases in late pregnancy(9/10)were detected respectively.4 of 36 cases cann’t be detected.Conclusion Fluorescent multiplex PCR can be used for amplification of male and female fetal STRs in maternal plasma to obtain genetic information,which may have implication for non-invasive prenatal diagnosis of certain hereditary diseases independent of the fetal sex.