Scalable and automated CRISPR-based strain engineering using droplet microfluidics

Abstract

We present a droplet-based microfluidic system that enables CRISPR-based gene editing and high-throughput screening on a chip. The microfluidic device contains a 10 × 10 element array, and each element contains sets of electrodes for two electric field-actuated operations: electrowetting for merging droplets to mix reagents and electroporation for transformation. This device can perform up to 100 genetic modification reactions in parallel, providing a scalable platform for generating the large number of engineered strains required for the combinatorial optimization of genetic pathways and predictable bioengineering. We demonstrate the system’s capabilities through the CRISPR-based engineering of two test cases: (1) disruption of the function of the enzyme galactokinase (galK) in E. coli and (2) targeted engineering of the glutamine synthetase gene (glnA) and the blue-pigment synthetase gene (bpsA) to improve indigoidine production in E. coli.

Perspectives for self-driving labs in synthetic biology

“…Microfluidics offer the opportunity to provide this seamless integration by encapsulating cells and reagents into droplets, and manipulating them precisely. Indeed, microfluidic platforms have been proposed for miniaturization of biological reactions, including DNA synthesis and assembly [40], transformation [41,42], cell-free expression [43], and phenotypic screening by fluorescence [44] and mass spectrometry [45]. Truly disruptive functionalities can be achieved by combining these capabilities with new developments in molecular sensors embedded on semiconductor chips [46], wireless optically activated microscopic sensors [47], monitoring of free radicals through fluorescent nanodiamonds [48], metabolic modulation through optogenetics [49], or manipulation of cells with light [50].…”

Perspectives in growth production trade-off in microbial bioproduction

“…Despite the essentiality of media optimization it remains ad hoc at least in the reported literature 73 and few examples of high throughput data driven media optimization exist 74,75 . One can anticipate that with improvements in automation and data driven approaches, and the ability to examine configurations in high throughput (e.g.Via microfluidics 76 ) this aspect will see a lot of improvements too. A consilience of metabolic, host engineering approaches with optimization of production medium would enable ideal growth-production ratios that are stable (and thus predictable) across scales.…”

“…74,75 One can anticipate that with improvements in automation and data-driven approaches and the ability to examine configurations in high throughput ( e.g. , via microfluidics 76 ), this aspect will also witness a lot of improvements. The consilience of metabolic and host engineering approaches with the optimization of production medium will enable ideal growth-production ratios that are stable (and thus predictable) across scales.…”

Biological research and self-driving labs in deep space supported by artificial intelligence

“…In the past decade, advances in several research areas have made such self-driving labs possible on Earth [88][89][90][91][92][93] . We now have the ability to automate many biological processes using state-of-the-art microfluidics chips for optics, imaging and robotics [94][95][96][97][98] .…”