Virtual Microfluidics for digital quantification and single-cell sequencing

Interest in highly parallelized analysis of single molecules and single cells is growing rapidly. Here we develop hydrogel-based virtual microfluidics as a simple alternative to complex engineered microfluidic systems for the compartmentalization of nucleic acid amplification reactions. We applied digital multiple displacement amplification (dMDA) to purified DNA templates, cultured bacterial cells, and human microbiome samples in the virtual microfluidics system and demonstrated recovery and whole-genome sequencing of single-cell MDA products. Our results from control samples showed excellent coverage uniformity and markedly reduced chimerism compared with single-cell data obtained from conventional liquid MDA reactions. We also demonstrate the applicability of the hydrogel method for genomic studies of naturally occurring microbes in human microbiome samples. The virtual microfluidics approach is a simple and robust method that will enable many laboratories to perform single-cell genomic analyses.