Wu Haitao, Ding Chenhui, Shen Xiaoting, Wang Jing, Li Rong, Cai Bing, Xu Yanwen, Zhong Yiping, Zhou Canquan
From the Reproductive Medicine Center, First Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong, China, Guangdong Provincial Key Laboratory of Reproductive Medicine.
Medicine (Baltimore). 2015 Mar;94(12):e669. doi: 10.1097/MD.0000000000000669.
To develop a noninvasive medium-based preimplantation genetic diagnosis (PGD) test for α-thalassemias-SEA. The embryos of α-thalassemia-SEA carriers undergoing in vitro fertilization (IVF) were cultured. Single cells were biopsied from blastomeres and subjected to fluorescent gap polymerase chain reaction (PCR) analysis; the spent culture media that contained embryo genomic DNA and corresponding blastocysts as verification were subjected to quantitative-PCR (Q-PCR) detection of α-thalassemia-SEA. The diagnosis efficiency and allele dropout (ADO) ratio were calculated, and the cell-free DNA concentration was quantitatively assessed in the culture medium. The diagnosis efficiency of medium-based α-thalassemias-SEA detection significantly increased compared with that of biopsy-based fluorescent gap PCR analysis (88.6% vs 82.1%, P < 0.05). There is no significant difference regarding ADO ratio between them. The optimal time for medium-based α-thalassemias-SEA detection is Day 5 (D5) following IVF. Medium-based α-thalassemias-SEA detection could represent a novel, quick, and noninvasive approach for carriers to undergo IVF and PGD.
开发一种用于α地中海贫血东南亚型（α-thalassemias-SEA）的基于培养基的非侵入性植入前基因诊断（PGD）检测方法。对接受体外受精（IVF）的α地中海贫血东南亚型携带者的胚胎进行培养。从卵裂球中活检单个细胞，并进行荧光缺口聚合酶链反应（PCR）分析；对含有胚胎基因组DNA的废弃培养基以及相应囊胚作为验证，进行α地中海贫血东南亚型的定量PCR（Q-PCR）检测。计算诊断效率和等位基因脱扣（ADO）率，并对培养基中的游离DNA浓度进行定量评估。与基于活检的荧光缺口PCR分析相比，基于培养基的α地中海贫血东南亚型检测的诊断效率显著提高（88.6%对82.1%，P<0.05）。两者之间的ADO率无显著差异。基于培养基的α地中海贫血东南亚型检测的最佳时间是IVF后的第5天（D5）。基于培养基的α地中海贫血东南亚型检测可为携带者进行IVF和PGD提供一种新的、快速且非侵入性的方法。
