Fluorescence-activated droplet sorting of lipolytic microorganisms using a compact optical system

Abstract

Lipases are ubiquitous enzymes of great physiological significance that have been used extensively in multiple industries. Environmental microorganisms are a major source for the discovery of novel lipases with high catalytic efficiency and selectivity. However, current plate-based screening of lipase-producing strains is time consuming, labour intensive and inefficient. In this study, we developed an ultra-high throughput screening pipeline for lipase-producing strains based on fluorescence-activated droplet sorting (FADS) using a compact optical system that could be easily set up in an alignment-free manner. The pipeline includes droplet generation, droplet incubation, picoinjection of the fluorescence probe, and sorting of droplets with a throughput of 2 × 10 drops per h. We applied the pipeline to screen samples collected from different locations, including sediments from a hot spring in Tibet, soils from the Zoige wetland, contaminated soils from an abandoned oilfield, and a Chinese Daqu starter. In total, we obtained 47 lipase-producing bacterial strains belonging to seven genera, including Staphylococcus, Bacillus, Enterobacter, Serratia, Prolinoborus, Acinetobacter, and Leclercia. We believe that this FADS-based pipeline could be extended to screen various enzymes from the environment, and may find wide applications in breeding of industrial microorganisms.

Characterisation of physicochemical properties, flavour components and microbial community in Chinese Guojing roasted sesame-like flavourDaqu

“…Maifreni et al reported an amino acid decarboxylase activity from Enterobacter from Montasio cheese (29). Enterobacter have been found to produce lipase, which plays an important role in the formation of esters in Baijiu, by an ultra high-throughput screening (30). A relatively strong correlation was found between saccharifying and liquefied enzyme activities and Enterobacter (31).…”

Comprehensive analysis of different grades of roasted-sesame-like flavored Daqu

“…[20] For example, Bacillus, Enterobacter, and Staphylococcus produce lipase. [40] Pediococcus and Weissella are lactic acid bacteria and produce not only lactic acid as a major end-product of carbohydrate fermentation but also provide substrates for esterification by yeasts. [27] The RDA results for the eukaryotes (Figure 4b) indicated that there were significant positive correlations between Pichia, Rasamsonia, Rhizomucor, Saccharomyces, and unidentified Eugregarinorida in sample A and between Absidia, Lichtheimia, Rhizopus, and Thermomucor in sample C. The RDA results indicated that Lichtheimia, Rhizopus, and Thermomucor, positively correlated with the saccharification activity and Pichia, Rhizopus, Saccharomyces, and Thermomucor positively correlated with the esterification activity.…”

Fluorescent nucleic acid probe in droplets for bacterial sorting (FNAP-sort) as a high-throughput screening method for environmental bacteria with various growth rates

“…Despite its considerable potential, only a few droplet microfluidics-based platforms for the cultivation of environmental microorganisms have been reported to date [19, 20]. The lack of simple detection and sorting methods for droplets containing bacteria hampers the widespread use of droplet microfluidics techniques for this purpose.…”