High-Throughput Picodroplet-Based Analysis of Biosynthetic Libraries

Biosynthesis of high value chemicals by engineered microorganisms is an application of synthetic biology that offers both economic and environmental advantages. This application is increasing the need for high-throughput screening tools that can facilitate the detection of the best performance among a library of designed microbes. For this reason we are developing a high-throughput, miniaturised Mass Spectrometry (MS) tool for profiling synthetic designed libraries ⁽¹⁾.

Combining microfluidics based picodroplet technology for cell encapsulation and sorting together with Mass Spectroscopy we aim to rapidly screen, identify and retrieve the best cell “hits” among synthetic metabolic pathway libraries. Based on this novel approach we will be able to determine which construct has the genetic combination that gives the best biosynthesis performance. To test this new tool we have designed three libraries of two synthetic metabolic pathways using molecular engineering techniques ^(2,3,4).

We chose two previously described synthetic pathways to produce non-natural amino acids ^(5,6) and focus on improving their level of expression. Various strategies have been explored such as the use of homologue genes from other organisms, varying the DNA copy number, transcription levels or translation activity. Then, using a pioneering picodroplet-based technology ⁽⁷⁾that enables not only for the testing of up to 200,000 samples per day by MS, using miniaturised input volumes (400-700 pL) but also for retrieving identified ‘hits’ in a reproducible manner, we will select single cells, analyse their production of this non-natural amino acids and finally select and recover the best performing clones among the different profiles obtained for further studies.

This will enable new scientific breakthroughs, higher throughputs, lower screening costs, shorten design-build-test cycle and thus, be of interest to the current MS user base in the synthetic biology market and other sectors.

1. Joint project between Imperial College London and Sphere Fluidics Limited is jointly supported by a Technology Strategy Board Grant and the BBSRC.
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