Microfluidic Assembly of Synthetic Ecologies for Drug Discovery (MASEDD)
Abstract
We have used our expertise in experimental evolution and adaptation to develop Microfluidic Assembly of Synthetic Ecologies for Drug Discovery or MASEDD as a novel approach to the discovery of new antimicrobial leads and antimicrobial strategies. MASEDD drives Streptomyces toward production of new antimicrobials by linking Streptomyces reproductive success to the ability to kill clinically relevant pathogens. We have built and optimized microfluidic devices for the rapid generation of water-in-oil droplets to co-encapsulate two strains: a Predator and Prey. The Predator strain (S. venezuelae) has been chromosomally labeled with a gene encoding red fluorescent protein (RFP) and chemically mutagenized to increase diversity. The Prey strain (Pseudomonas aeruginosa) is labelled with a green fluorescent protein (GFP) encoding plasmid. Growth conditions have been optimized to achieve comparable growth rates of both strains. Following co-encapsulation, we use Fluorescence Assisted Droplet Sorting (FADS) to enrich for successful S. venezuelae lineages that have the ability to inhibit the Prey (pathogen). In addition to the expensive microscope-based FADS device, we have constructed a bench-top device which is cost effective, allowing us to have multiple FADS devices running simultaneously to allow sorting of multiple replicate populations. Following droplet sorting, the challenge was to separate the predator strain from the prey cells for the next round of encapsulation. We have been able to devise a simple solution for this problem by taking advantage of the extensive branching morphology of Streptomycetes. We find that using a 20 micrometers pore size filter allows effective separation of the homogenously growing planktonic Prey cells from the Predator.
Document Details
- Document Type
- Technical Report
- Publication Date
- Oct 01, 2020
- Accession Number
- AD1120988
Entities
People
- Yousif Shamoo
Organizations
- Rice University