Department of Medical Genetics, School of Basic Medical Sciences, Southern Medical University, Guangzhou, China.
Department of Birth Health and Heredity, Liuzhou Municipal Maternity and Child Healthcare Hospital, Guangxi, Liuzhou, China.
J Hum Genet. 2018 Apr;63(4):407-416. doi: 10.1038/s10038-018-0411-5. Epub 2018 Feb 13.
In this study, we aimed to assess the performance of two whole-genome amplification methods, multiple displacement amplification (MDA), and multiple annealing and looping-based amplification cycle (MALBAC), for β-thalassemia genotyping and single-nucleotide polymorphism (SNP)/copy-number variant (CNV) detection using two DNA sequencing assays. We collected peripheral blood, cell lines, and discarded embryos, and carried out MALBAC and MDA on single-cell and five-cell samples. We detected and statistically analyzed differences in the amplification efficiency, positive predictive value, sensitivity, allele dropout (ADO) rate, SNPs, and CV values between the two methods. Through Sanger sequencing at the single-cell and five-cell levels, we showed that both the amplification rate and ADO rate of MDA were better than those using MALBAC, and the sensitivity and positive predictive value obtained from MDA were higher than those from MALBAC for β-thalassemia genotyping. Using next-generation sequencing (NGS) at the single-cell level, we confirmed that MDA has better properties than MALBAC for SNP detection. However, MALBAC was more stable and homogeneous than MDA using low-depth NGS at the single-cell level for CNV detection. We conclude that MALBAC is the better option for CNV detection, while MDA is better suited for SNV detection.
在这项研究中，我们旨在评估两种全基因组扩增方法——多重置换扩增（MDA）和多次退火和环化扩增循环（MALBAC）——在使用两种 DNA 测序检测方法进行β-地中海贫血基因分型和单核苷酸多态性（SNP）/拷贝数变异（CNV）检测中的性能。我们收集了外周血、细胞系和废弃胚胎，并对单细胞和五细胞样本进行了 MALBAC 和 MDA 扩增。我们检测并统计分析了两种方法在扩增效率、阳性预测值、灵敏度、等位基因缺失（ADO）率、SNP 和 CV 值方面的差异。通过单细胞和五细胞水平的 Sanger 测序，我们表明 MDA 的扩增率和 ADO 率均优于 MALBAC，并且 MDA 获得的灵敏度和阳性预测值均高于 MALBAC 用于β-地中海贫血基因分型。通过单细胞水平的下一代测序（NGS），我们证实 MDA 在 SNP 检测方面优于 MALBAC。然而，对于单细胞水平的低深度 NGS 进行 CNV 检测，MALBAC 比 MDA 更稳定和均匀。我们得出结论，MALBAC 是 CNV 检测的更好选择，而 MDA 更适合 SNP 检测。
