| Summary: | <i>Haematococcus pluvialis</i> is a microalgae actively studied for the production of natural astaxanthin, which is a powerful antioxidant for human application. However, it is economically disadvantageous for commercialization owing to the low productivity of astaxanthin. This study reports an effective screening strategy using the negative phototaxis of the <i>H. pluvialis</i> to attain the mutants having high astaxanthin production. A polydimethylsiloxane (PDMS)-based microfluidic device irradiated with a specific light was developed to efficiently figure out the phototactic response of <i>H. pluvialis</i>. The partial photosynthesis deficient (PP) mutant (negative control) showed a 0.78-fold decreased cellular response to blue light compared to the wild type, demonstrating the positive relationship between the photosynthetic efficiency and the phototaxis. Based on this relationship, the <i>Haematococcus</i> mutants showing photosensitivity to blue light were selected from the 10,000 random mutant libraries. The M1 strain attained from the phototaxis-based screening showed 1.17-fold improved growth rate and 1.26-fold increases in astaxanthin production (55.12 ± 4.12 mg g<sup>−1</sup>) in the 100 L photo-bioreactor compared to the wild type. This study provides an effective selection tool for industrial application of the <i>H. pluvialis</i> with improved astaxanthin productivity.
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