基于父源差异SNP位点的等位基因特异性实时荧光PCR方法建立及其在.docVIP

基于父源差异SNP位点的等位基因特异性实时荧光PCR方法建立及其在α0-地中海贫血中的无创产前诊断中的应用 刘彦慧1,吴亚敏2, 石少权3,娄季武1, 马秋林1, 何怡1, 徐婉芳1, 林洋洋1 ,周万军4（523700 广东 东莞，东莞市妇幼保健院产前诊断中心1, 523220广东 东莞，东华医院中心实验室2, 519000广东珠海，中山大学第五附属医院妇产科3, 510515广州，南方医科大学基础医学院医学遗传学教研室4） [摘要] 目的 建立并评价1种纯合α0-地中海贫血排除性无创产前诊断的等位基因特异性实时荧光PCR技术(AS-qPCR)。方法 用PCR微测序技术，检测SEA高风险夫妇基因缺失区内9个SNP以确定双亲差异位点；然后以AS-qPCR技术检测孕妇血浆中父源差异SNP位点，判断父方是否将正常单倍型遗传给胎儿。结果 167个家系中，160个家系找到1个或多个有双亲差异的SNP位点，检测率约为98.16%；94例检测到父源性正常等位基因SNP而免于创伤性产前诊断，检出率57.67%；所有家系的绒毛或羊水基因检测结果与本方法的符合率为100%。 结论 采用AS-qPCR检测孕妇血浆游离DNA中双亲差异SNP位点的方法，在孕早期即可进行纯合α0-地中海贫血的排除性无创产前诊断，可使约50%高风险孕产妇免于创伤性产前诊断。 [关键词] α-地中海贫血；无创产前诊断；游离DNA；基因缺失 [中图法分类号] [文献标志码] A The development and application of allele-specific real-time fluorescence PCR for homozygous ?0-thalassemia non-invasive prenatal diagnosis based on paternal different SNP Liu Yanhui1, 　Wu Yamin2,　Shi Shaoquan3,　Lou Jiwu1，Ma Qiulin1,　He Yi1,　Xu Wanfang1,　Lin Yangyang1,　Zhou Wanjun4*　（1 prenatal diagnosis center, Maternal and Child Health Hospital of DongGuan, GuangDong, 523700,china. 2 Center Laboratory, Donghua Hospital, Dongguan,GuangDong, 523220,China. 3 The Fifth Affiliated Hospital, Sun Yat-Sen University, ZhuHai, GuangDong, 519000, China. 4 Department of Genetics，Southern Medical University, GuangZhou, GuangDong, 510515, China.） [Abstract] Objective To establish and evaluate an allele-specific real-time fluorescence PCR (AS-QPCR) assay for homozygous a0-thalassemia exclusive non-invasive prenatal diagnosis. Methods Among 9 SNPs within the SEA deletion range, the different SNPs between parents were determined with PCR micro-sequencing technology. The paternal SNP was detect in maternal plasma DNA with AS-qPCR method to determine whether the paternal normal haplotype inherited to the fetus. Results 160 of 167 families were found have one or more parents-difference SNPs with the detection rates over 98.16%. The paternal normal SNPs were detected in 94 maternal plasma DNA and then they avoid invasive prenatal di