Uniform and accurate single-cell sequencing based on emulsion whole-genome amplification

Significance Uniform and accurate single-cell whole-genome amplification is important when starting material is limited and precious. We develop an emulsion-based amplification method that can suppress the amplification bias to detect high-resolution copy number variations of a single cell, and to simultaneously detect the single-nucleotide variations with high accuracy. This approach is compatible with various amplification protocols including the widely used multiple displacement amplification, which has been demonstrated in this paper. 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 (105) 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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