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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。
Objective To establish and evaluate an allele-specific real-time fluorescent PCR technique (AS-qPCR) for the detection of a homozygous α0-thalassemia noninvasive prenatal diagnosis. Methods PCR micro-sequencing technology was used to detect nine SNPs in the gene-deleted region of SEA high-risk couples to determine the differences between the two parents; then AS-qPCR was used to detect the paternal SNPs in plasma of pregnant women, Genetics to the fetus. Results One hundred and sixty-six pedigrees found one or more SNPs with two parents, the detection rate was 98.16%; 94 cases were detected SNPs of paternal normal allele without being diagnosed by traumatic prenatal diagnosis, The detection rate was 57.67%. The detection rate of villus or amniotic fluid in all pedigrees was 100% in accordance with the method. Conclusions AS-qPCR can be used to detect the difference between the two parents in the plasma free DNA of pregnant women. In the early stage of pregnancy, the exclusion of homozygous α0-thalassemia can be performed without prenatal diagnosis, and about 50% of the high-risk pregnant women can be protected from Traumatic prenatal diagnosis.