High-throughput screening of filamentous fungi using nanoliter-range droplet-based microfluidics.
Filamentous fungi are an extremely important source of industrial enzymes because of their capacity to secrete large quantities of proteins. Currently, functional screening of fungi is associated with low throughput and high costs, which severely limits the discovery of novel enzymatic activities and better production strains. Here, we describe a nanoliter-range droplet-based microfluidic system specially adapted for the high-throughput sceening (HTS) of large filamentous fungi libraries for secreted enzyme activities. The platform allowed (i) compartmentalization of single spores in ~10 nl droplets, (ii) germination and mycelium growth and (iii) high-throughput sorting of fungi based on enzymatic activity. A 10(4) clone UV-mutated library of Aspergillus niger was screened based on α-amylase activity in just 90 minutes. Active clones were enriched 196-fold after a single round of microfluidic HTS. The platform is a powerful tool for the development of new production strains with low cost, space and time footprint and should bring enormous benefit for improving the viability of biotechnological processes.
École Supérieure de Physique et de Chimie Industrielles de la Ville de Paris (ESPCI Paris), CNRS UMR 8231, 10, rue Vauquelin, 75231 Paris Cedex 05, France.
Thomas
T
Beneyton
École Supérieure de Physique et de Chimie Industrielles de la Ville de Paris (ESPCI Paris), CNRS UMR 8231, 10, rue Vauquelin, 75231 Paris Cedex 05, France.|Ets J. Soufflet/CRIS-OSIRIS, Quai Sarrail, BP12, 10400 Nogent-sur-Seine, France.
I Putu Mahendra
IP
Wijaya
Ets J. Soufflet/CRIS-OSIRIS, Quai Sarrail, BP12, 10400 Nogent-sur-Seine, France.
Prexilia
P
Postros
Ets J. Soufflet/CRIS-OSIRIS, Quai Sarrail, BP12, 10400 Nogent-sur-Seine, France.
Majdi
M
Najah
Ets J. Soufflet/CRIS-OSIRIS, Quai Sarrail, BP12, 10400 Nogent-sur-Seine, France.
Pascal
P
Leblond
Ets J. Soufflet/CRIS-OSIRIS, Quai Sarrail, BP12, 10400 Nogent-sur-Seine, France.
Angélique
A
Couvent
Ets J. Soufflet/CRIS-OSIRIS, Quai Sarrail, BP12, 10400 Nogent-sur-Seine, France.
Estelle
E
Mayot
École Supérieure de Physique et de Chimie Industrielles de la Ville de Paris (ESPCI Paris), CNRS UMR 8231, 10, rue Vauquelin, 75231 Paris Cedex 05, France.
Andrew D
AD
Griffiths
Ets J. Soufflet/CRIS-OSIRIS, Quai Sarrail, BP12, 10400 Nogent-sur-Seine, France.
Antoine
A
Drevelle
D001234:Aspergillus niger / Q000201:enzymology / Q000235:genetics / Q000302:isolation & purification*; D057166:High-Throughput Screening Assays / Q000379:methods; D046210:Microfluidic Analytical Techniques / Q000379:methods*; D009154:Mutation; D000516:alpha-Amylases / Q000235:genetics / Q000378:metabolism*
D000516:alpha-Amylases