Photosynthetic organisms evolved phototaxis to find optimal light condition for photosynthesis. Here we report high-throughput screening system using phototaxis of Chlamydomonas reinhardtii, for rapid isolation of strains with enhanced photosynthetic efficiencies in microfluidic system. We proved the correlation between phototaxis and PSII efficiency by quantitative analysis of phototaxis. We enriched the strains with improved PSII efficiency by isolating cells showing fast phototactic response from 10,000 mutants. Among 147 strains isolated after screening, 94.6% showed enhanced growth over the parental strain. Two mutants showed much improved performances with up to 1.9- and 8.1-fold increases in growth and lipid production, respectively. Our approach provides a powerful screening tool for rapid improvement of microalgae to enhance photosynthetic efficiency.