Noda Yoshiteru, Kato Takema, Kato Asuka, Nishizawa Haruki, Miyazaki Jun, Ito Mayuko, Terasawa Sumire, Sekiya Takao, Fujii Takuma, Kurahashi Hiroki
Department of Obstetrics and Gynecology, Fujita Health University School of Medicine, Toyoake, Japan.
Division of Molecular Genetics, Institute for Comprehensive Medical Science, Fujita Health University, Toyoake, Japan.
Congenit Anom (Kyoto). 2019 May;59(3):88-92. doi: 10.1111/cga.12302. Epub 2018 Jul 6.
Examination of maternal plasma cell-free DNA (cfDNA) for noninvasive prenatal testing for fetal trisomy is a highly effective method for pregnant women at high risk. This can be also applied to fetal gender determination in female carriers of severe X-linked disease. Polymerase chain reaction (PCR) analysis is a relatively simpler and less expensive method of detecting Y chromosome-specific repeats (Y-specific PCR; YSP), but is limited by the risk of false-negative results. To address this, we have developed a combined strategy incorporating YSP and an estimation of the fetal DNA fraction. Multiplex PCR for 30 single nucleotide polymorphism (SNP) loci selected by high heterozygosity enables the robust detection of the fetal DNA fraction in cfDNA. The cfDNA sample is first subjected to YSP. When the YSP result is positive, the fetus is male and invasive testing for an X-linked mutation is then required. When the YSP result is negative, the cfDNA sample is analyzed using multiplex PCR. If fetal DNA is then found in the cfDNA, invasive testing is not then required. If the multiplex PCR analysis of cfDNA is negative for fetal DNA, the fetal gender cannot be determined and invasive testing is still required. Our technique provides a potentially effective procedure that can help to avoid unnecessary invasive prenatal testing in some female carriers of severe X-linked disease.
检测孕妇血浆游离DNA（cfDNA）用于胎儿三体非侵入性产前检测，对于高危孕妇是一种高效的方法。这也可应用于严重X连锁疾病女性携带者的胎儿性别鉴定。聚合酶链反应（PCR）分析是检测Y染色体特异性重复序列（Y特异性PCR；YSP）相对简单且成本较低的方法，但受假阴性结果风险的限制。为解决此问题，我们开发了一种结合YSP和胎儿DNA分数估计的联合策略。针对通过高杂合性选择的30个单核苷酸多态性（SNP）位点进行多重PCR，能够可靠地检测cfDNA中的胎儿DNA分数。首先对cfDNA样本进行YSP检测。当YSP结果为阳性时，胎儿为男性，然后需要进行X连锁突变的侵入性检测。当YSP结果为阴性时，使用多重PCR分析cfDNA样本。如果随后在cfDNA中发现胎儿DNA，则不需要进行侵入性检测。如果cfDNA的多重PCR分析对胎儿DNA呈阴性，则无法确定胎儿性别，仍需要进行侵入性检测。我们的技术提供了一种潜在有效的程序，有助于避免一些严重X连锁疾病女性携带者进行不必要的侵入性产前检测。
