Journal
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Volume 112, Issue 38, Pages 11923-11928Publisher
NATL ACAD SCIENCES
DOI: 10.1073/pnas.1513988112
Keywords
single cell; whole-genome amplification; sequencing; microfluidics; emulsion
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Funding
- National Natural Science Foundation of China [21327808, 91313302, 21222501]
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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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