A Microfluidic Platform for the Discovery and Functional Annotation of Metagenomic Enzymes

It is believed that the enzymatic machinery of microorganisms can be harnessed to produce sustainable bioenergy as well as biosequestration of carbon. However, the ability to engineer organisms is contingent on being able to both understand and take advantage of these functions of interest. Recent advances in high-throughput sequencing have dramatically increased the speed of progress in these areas. However, the task of connecting this genomic scale information with catabolic functions is progressing relatively slowly. Annotating genes by quickly connecting large-scale functions of interest to their genetic circuits and catabolic pathways is a difficult task that requires new tools that do not currently exist. We propose a high-throughput, microfluidic platform technology to identify genes and metabolic networks involved in producing enzymes with specific activity. This technology operates with a three-tiered approach. First a metagenomics library of genetic elements from soil microbiota is created and transferred into an Escherichia coli host. Cells that affect selected enzyme substrates are identified and isolated within the microfluidic chip. Finally, the expression vector of the selected cells will be sequenced. This raw data can then be subjected to bioinformatic analysis to trace the genetic elements back to their microbial origin and annotate them with the detected enzymatic activity. In summary, the concept this technology is to take a enzyme substrate and rapidly find metagenomic sequences involved in its conversion. In contrast with a horizontal approach where the function of all genetic elements of a single organism would be annotated, this is a vertical approach to the identification and annotation of all enzymes coding genetic elements in the library comprising numerous species acting on a specific substrate. Our method allows us to quickly sample the enormous functional reservoir of novel genes from a metagenome for applications in bioenergy, biosequestration, bioremediation and beyond.