Directed Evolution in Drops: Molecular Aspects and Applications

Abstract

The process of optimizing the properties of biological molecules is paramount for many industrial and medical applications. Directed evolution is a powerful technique for modifying and improving biomolecules such as proteins or nucleic acids (DNA or RNA). Mimicking the mechanism of natural evolution, one can enhance a desired property by applying a suitable selection pressure and sorting improved variants. Droplet-based microfluidic systems offer a high-throughput solution to this approach by helping to overcome the limiting screening steps and allowing the analysis of variants within increasingly complex libraries. Here, we review cases where successful evolution of biomolecules was achieved using droplet-based microfluidics, focusing on the molecular processes involved and the incorporation of microfluidics to the workflow. We highlight the advantages and limitations of these microfluidic systems compared to low-throughput methods and show how the integration of these systems into directed evolution workflows can open new avenues to discover or improve biomolecules according to user-defined conditions.

FACS-Sortable Triple Emulsion Picoreactors for Screening Reactions in Biphasic Environments

“…Proteins can be expressed in situ within triple emulsion picoreactors. Enzymes catalysts are particularly amenable to high-throughput screening strategies when produced via in vitro cell-free protein synthesis within an aqueous phase 40,41 . To test whether our triple emulsion picoreactors can be used to express and screen enzyme catalysts, we first tested compatibility of in vitro transcription/translation (IVTT) with reagents required to form and stabilize droplets (Figure 4A).…”

The Importance of Dean Flow in Microfluidic Nanoparticle Synthesis: A ZIF‐8 Case Study

“…The microscale tubing ensures fast and uniform heat transfer, and the microfluidic setup can be easily automated. [7][8][9][10] Fields such as biosensing, [11] bio(medical)engineering, [12][13][14][15][16] microbiology, [17,18] organic synthesis, [19,20] inorganic chemistry, [21] catalysis, [22] and many others rely on microfluidic synthesis, so advances in this technology directly impact numerous applications.…”

Crafting Genetic Diversity: Unlocking the Potential of Protein Evolution

“…The combination of a largersized and higher-quality library enables more efficient exploration of the protein space, particularly when the exploration is not constrained by screening/selection capacity. This review deliberately omits the discussion of screening or selection as it has been extensively covered in our textbook [5] and several recent, excellent reviews [66][67][68][69][70][71]. Readers are encouraged to consult these references for a comprehensive understanding of this aspect.…”