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RNA-aptamers-in-droplets (RAPID) high-throughput screening for secretory phenotypes.

  • Author(s): Abatemarco, Joseph
  • Sarhan, Maen
  • Wagner, James
  • Lin, Jyun-Liang
  • Liu, Leqian
  • Hassouneh, Wafa
  • Yuan, Shuo-Fu
  • Alper, Hal
  • Abate, Adam
  • et al.

Synthetic biology and metabolic engineering seek to re-engineer microbes into living foundries for the production of high value chemicals. Through a design-build-test cycle paradigm, massive libraries of genetically engineered microbes can be constructed and tested for metabolite overproduction and secretion. However, library generation capacity outpaces the rate of high-throughput testing and screening. Well plate assays are flexible but with limited throughput, whereas droplet microfluidic techniques are ultrahigh-throughput but require a custom assay for each target. Here we present RNA-aptamers-in-droplets (RAPID), a method that greatly expands the generality of ultrahigh-throughput microfluidic screening. Using aptamers, we transduce extracellular product titer into fluorescence, allowing ultrahigh-throughput screening of millions of variants. We demonstrate the RAPID approach by enhancing production of tyrosine and secretion of a recombinant protein in Saccharomyces cerevisiae by up to 28- and 3-fold, respectively. Aptamers-in-droplets affords a general approach for evolving microbes to synthesize and secrete value-added chemicals.Screening libraries of genetically engineered microbes for secreted products is limited by the available assay throughput. Here the authors combine aptamer-based fluorescent detection with droplet microfluidics to achieve high throughput screening of yeast strains engineered for enhanced tyrosine or streptavidin production.

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