Microfluidic high-throughput selection of microalgal strains with superior photosynthetic productivity using competitive phototaxis

Abstract

Microalgae possess great potential as a source of sustainable energy, but the intrinsic inefficiency of photosynthesis is a major challenge to realize this potential. Photosynthetic organisms evolved phototaxis to find optimal light condition for photosynthesis. Here we report a microfluidic screening using competitive phototaxis of the model alga, Chlamydomonas reinhardtii, for rapid isolation of strains with improved photosynthetic efficiencies. We demonstrated strong relationship between phototaxis and photosynthetic efficiency by quantitative analysis of phototactic response at the single-cell level using a microfluidic system. Based on this positive relationship, we enriched the strains with improved photosynthetic efficiency by isolating cells showing fast phototactic responses from a mixture of 10,000 mutants, thereby greatly improving selection efficiency over 8 fold. Among 147 strains isolated after screening, 94.6% showed improved photoautotrophic growth over the parental strain. Two mutants showed much improved performances with up to 1.9- and 8.1-fold increases in photoautotrophic cell growth and lipid production, respectively, a substantial improvement over previous approaches. We identified candidate genes that might be responsible for fast phototactic response and improved photosynthesis, which can be useful target for further strain engineering. Our approach provides a powerful screening tool for rapid improvement of microalgal strains to enhance photosynthetic productivity.

Quantitative analysis of the chemotaxis of a green alga, Chlamydomonas reinhardtii, to bicarbonate using diffusion-based microfluidic device

“…After the linear gradient in the channel was achieved, the TP medium in the cell reservoir was carefully removed, and 35 ll of the cell suspension was injected. In case of chemotaxis assay using NH 4 Cl, the harvested cells were resuspended in nitrogen-deprived TP medium (TPÀN medium, TP medium without NH 4 Cl) at an OD 800 of 1.0. The TPÀN medium was also used as the basis of NH 4 Cl solution with different concentrations (10, 50, and 100 mM) for the chemotaxis analysis.…”

“…In case of chemotaxis assay using NH 4 Cl, the harvested cells were resuspended in nitrogen-deprived TP medium (TPÀN medium, TP medium without NH 4 Cl) at an OD 800 of 1.0. The TPÀN medium was also used as the basis of NH 4 Cl solution with different concentrations (10, 50, and 100 mM) for the chemotaxis analysis. To prevent unintended phototaxis-involved cell movement, the microfluidic device was placed in dark during the assay period.…”

“…The number of cells in the observation region was determined through the manual counting and the use of software (ImageJ) to minimize errors in counting number of cells in the observation chamber. All reagents, including CoCl 2 , NaCl, and NH 4 Cl, were purchased from Sigma-Aldrich. The wild-type cells were grown to exponential phase under the low CO 2 and the high CO 2 conditions.…”

“…We validated the microfluidic device by analyzing chemotactic responses of C. reinhardtii to previously known chemoattractants (NH 4 Cl and CoCl 2 ) and the chemotactically neutral molecule (NaCl). 8,[10][11][12] It was reported that ammonium ion (NH 4 þ ) is a strong chemoattractant for C. reinhardtii, 8,10,12 but NaCl does not elicit any response (chemotactically neutral) 10 of the alga.…”

“…Despite the potential, it is still required to improve photosynthetic productivity for the economic viability of microalgal production systems. 4 A deeper understanding of algal physiology is crucial for the realization of biotechnological application of microalgae. 5,6 There have been a number of studies on the physiology related to photosynthesis, 7 circadian rhythm, 8 and tactic responses 9-12 using a unicellular motile green alga, Chlamydomonas reinhardtii, as a model organism of microalgae.…”