Droplet Tn-Seq combines microfluidics with Tn-Seq for identifying complex single-cell phenotypes

Derek Thibault, Paul A. Jensen, Stephen Wood, Christine Qabar, Stacie Clark, Mara G. Shainheit, Ralph R. Isberg, Tim van Opijnen
2019 Nature Communications  
While Tn-Seq is a powerful tool to determine genome-wide bacterial fitness in high-throughput, culturing transposon-mutant libraries in pools can mask community or other complex single-cell phenotypes. Droplet Tn-Seq (dTn-Seq) solves this problem by microfluidics facilitated encapsulation of individual transposon mutants into growth medium-in-oil droplets, thereby enabling isolated growth, free from the influence of the population. Here we describe and validate microfluidic chip design,
more » ... hip design, production, encapsulation, and dTn-Seq sample preparation. We determine that 1-3% of mutants in Streptococcus pneumoniae have a different fitness when grown in isolation and show how dTn-Seq can help identify leads for gene function, including those involved in hyper-competence, processing of alpha-1-acid glycoprotein, sensitivity against the human leukocyte elastase and microcolony formation. Additionally, we show dTn-Seq compatibility with microscopy, FACS and investigations of bacterial cell-to-cell and bacteria-host cell interactions. dTn-Seq reduces costs and retains the advantages of Tn-Seq, while expanding the method's original applicability.
doi:10.1038/s41467-019-13719-9 pmid:31844066 fatcat:u6gc5uhplbhb7mgi6hwbhucbfm