多重位移放大
拷贝数变化
基因组
生物
计算生物学
基因复制
低拷贝数
单细胞测序
人类基因组
DNA
基因组DNA
DNA测序
基因组学
遗传学
基因
表型
聚合酶链反应
外显子组测序
DNA提取
作者
Yusi Fu,Chunmei Li,Sijia Lü,Zhou Wei-ping,Fuchou Tang,X. Sunney Xie,Yanyi Huang
标识
DOI:10.1073/pnas.1513988112
摘要
Whole-genome amplification (WGA) for next-generation sequencing has seen wide applications in biology and medicine when characterization of the genome of a single cell is required. High uniformity and fidelity of WGA is needed to accurately determine genomic variations, such as copy number variations (CNVs) and single-nucleotide variations (SNVs). Prevailing WGA methods have been limited by fluctuation of the amplification yield along the genome, as well as false-positive and -negative errors for SNV identification. Here, we report emulsion WGA (eWGA) to overcome these problems. We divide single-cell genomic DNA into a large number (10(5)) of picoliter aqueous droplets in oil. Containing only a few DNA fragments, each droplet is led to reach saturation of DNA amplification before demulsification such that the differences in amplification gain among the fragments are minimized. We demonstrate the proof-of-principle of eWGA with multiple displacement amplification (MDA), a popular WGA method. This easy-to-operate approach enables simultaneous detection of CNVs and SNVs in an individual human cell, exhibiting significantly improved amplification evenness and accuracy.
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