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Droplet Microfluidics for Single-cell Genomics

Abstract

Microbial systems often exhibit staggering diversity, making the study of rare, interesting species challenging. Metagenomic analyses of mixed-cell populations are often dominated by the sequences of the most abundant organisms, while those of rare microbes are detected only at low levels, if at all. To overcome this, selective cultivation or fluorescence-activated cell sorting can be used to enrich for the target species prior to sequence analysis; however, since most microbes cannot be grown in the lab, cultivation strategies often fail, and while cell sorting requires techniques to uniquely label the cell type of interest, this not possible with uncultivable microbes. In this thesis, we introduce a strategy for the high-throughput targeted enrichment of microbial genomes, which we term PCR-activated cell sorting (PACS), and show that it enables the sequence-specific detection, sorting and recovery of microbial genomes. We then show that PACS can be applied to identify phage-host relationships by sorting bacteria based on infection by specific phages. PACS has many potential applications in microbiology, foremost among them the possibility to uncover the genetic makeup of microbial “dark matter”, in order for mankind to gain a greater understanding of these enigmatic creatures and the roles that they play in this world.

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