应用巢式PCR检测孕妇血浆中cffDNA RHD基因型的研究

发布时间：2018-06-30 09:44

本文选题：巢式PCR + 血浆游离胎儿DNA　；参考：《中国输血杂志》2016年08期

【摘要】：目的建立巢式PCR技术检测RhD阴性孕妇血浆中游离胎儿DNA(cffDNA)的RHD基因型,以预测胎儿RhD血型。方法采用QIAamp DNA Blood Mini Kit提取32例RhD阴性孕妇血浆游离DNA,针对RHD外显子7和10分别设计外侧、内侧2组特异性引物,巢式PCR方法检测孕妇血浆游离胎儿DNA的RHD型,测序验证PCR产物的序列特异性。结果孕妇血浆游离DNA经巢式PCR扩增后,有27例成功扩增出RHD外显子7、10特异性条带,5例未检测到RHD基因特异性扩增,32例中有30例胎儿RHD型与出生后血型相符,检测准确率为93.1%。结论采用巢式PCR技术检测RhD阴性孕妇血浆游离胎儿DNA来判定胎儿RHD型,具有良好的准确性、敏感性和特异性,为RhD新生儿溶血病的早期诊断提供了一种新的、可靠的检测手段。
[Abstract]:Objective to establish a nested PCR technique to detect the RHD genotypes of free fetal DNA (cffDNA) in the plasma of RhD negative pregnant women and to predict the fetal RhD blood group. Methods Plasma free DNA was extracted from 32 RhD-negative pregnant women by QIAamp DNA Blood Mini Kit. Two specific primers were designed for RHD exon 7 and 10, respectively. Nested PCR was used to detect the RHD type of free fetal DNA in maternal plasma. Sequence specificity of PCR products was verified by sequencing. Results in 27 cases of maternal plasma free DNA amplified by nested PCR, the RHD exon 710 specific band was successfully amplified in 5 cases. Of the 32 cases without RHD gene specific amplification, 30 cases of fetal RHD type corresponded to postnatal RHD blood type, and the detection accuracy was 93.1%. Conclusion Detection of free fetal DNA in plasma of RhD negative pregnant women by nested PCR has good accuracy, sensitivity and specificity, which provides a new and reliable method for the early diagnosis of RhD hemolytic disease.
【作者单位】：陕西省血液中心;南方医科大学南方医院;深圳市血液中心;
【分类号】：R714.5;R440

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