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α<sup>0</sup>-地中海贫血无创产前诊断中的应用-陆军军医大学学报（原第三军医大学学报）

文章信息/Info

Title:: Non-invasive prenatal diagnosis for homozygous α⁰-thalassemia based on paternal different SNP with allele-specific real-time fluorescence PCR: establishment and application

作者:: 刘彦慧; 吴亚敏; 石少权; 娄季武; 马秋林; 何怡; 徐婉芳; 林洋洋; 周万军; 东莞市妇幼保健院产前诊断中心；东华医院中心实验室；中山大学第五附属医院妇产科；南方医科大学基础医学院医学遗传学教研室

Author(s):: Liu Yanhui; Wu Yamin; Shi Shaoquan; Lou Jiwu; Ma Qiulin; He Yi; Xu Wanfang; Lin Yangyang; Zhou Wanjun; Center of Prenatal Diagnosis, Dongguan Maternal and Child Health Hospital, Dongguang, Guangdong Province, 523700; Laboratory Center, Donghua Hospital, Dongguan, Guangdong Province, 523220; Department of Gynecology and Obstetrics, Fifth Affiliated Hospital, Sun Yat-Sen University, Zhuhai, Guangdong Province, 519000; Department of Genetics, College of Basic Medical Sciences, Southern Medical University, Guangzhou, Guangdong Province, 510515, China

Keywords:: α-thalassemia; non-invasive prenatal diagnosis; free DNA; gene deletion

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。

Abstract:: Objective To establish and evaluate an allele-specific real-time fluorescence PCR (AS-qPCR) assay for non-invasive exclusive prenatal diagnosis of homozygous α⁰-thalassemia. 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 detected in maternal plasma DNA with AS-qPCR to determine whether the paternal normal haplotype had been inherited to the fetus. Results Among 167 families, 160 were found to have one or more parents-different SNPs with the detection rates over 98.16%. The paternal normal SNPs were detected in 94 maternal plasma DNA. In this way, they avoided invasive prenatal diagnosis with a detection rate of 57.67%. These results were 100% coincidence with the results of CVS or amniocentesis. Conclusion Using AS-qPCR to detect the parents-difference SNPs in maternal plasma DNA in the early stage of gestation, is a non-invasive exclusive prenatal diagnosis of homozygous α⁰-thalassemia. This method can make about 50% high-risk pregnant women to avoid invasive prenatal diagnosis.