High-throughput fluorescence-activated cell sorting for cell wall-deficient microalgal mutants screening

Bioavailability and extraction efficiency of the natural antioxidant astaxanthin from the unicellular green alga Haematococcus pluvialis have been limited by the rigid algal cell walls. A high-throughput fluorescence-activated cell sorting (FACS) pipeline was developed in this study with the aim to screen cell wall-deficient mutants. The fluorescein-conjugated lectin-Ricinus communis Agglutinin I (RCA(120)) was used as a marker for binding the sugar moieties of the algal cell walls, which enabled discriminating the algal strains that differed in the cell walls. By utilizing the established FACS pipeline, a cell wall-deficient mutant (i.e. mutant-264) with the significantly reduced RCA(120)-fluorescein intensity was screened from the mutant library. The total content of the cell wall-associated sugars was reduced by 43.9% in the mutant-264 as compared to that of the wild type, with significant decrease in mannose, mannitol and galactose. The dry weight, astaxanthin content and extraction efficiency were improved by 21.1%, 13.1% and 16.5%, respectively, in mutant-264 as compared to that of the wild type. This study not only demonstrated the feasibility of utilization of the FACS-based high-throughput pipeline for cell wall mutant screening, but also delivered a novel promising H. pluvialis strain with multiple improved traits for industrial applications.