Genome shuffling of the nonconventional yeast Pichia anomala for improved sugar alcohol production

Guoqiang Zhang, Yuping Lin, Xianni Qi, Lixian Wang, Peng He, Qinhong Wang, Yanhe Ma

Sugar alcohols have been widely applied in the fields of food and medicine owing to their unique properties. Compared to chemical production, microbial production of sugar alcohols has become attractive because of its environmentally friendly and sustainable characteristics. Our previous study identified the nonconventional yeast Pichia anomala TIB-x229 as a potential producer of sugar alcohols from glucose. To further improve strain performance, we combined genome shuffling with optimized high throughput screening methods for the directed improvement of nonconventional yeast and complex phenotypes. To accelerate strain improvement, a practical genome shuffling procedure was developed and successfully applied in the nonconventional yeast P. anomala to increase sugar alcohol production. Through two rounds of genome shuffling, an improved P. anomala isolate GS2-3 could produce 47.1 g/L total sugar alcohols from 100 g/L glucose, which was 32.3% higher than the original strain. In this process, a simple and accurate colorimetric assay was optimized and used for high throughput screening of sugar alcohol-producing strains. Moreover, a fluorescence-activated cell sorting method was developed to efficiently screen protoplast fusions for genome shuffling of nonconventional yeast. An efficient genome shuffling procedure was developed and applied to enhance the sugar alcohol production of the nonconventional yeast P. anomala. Our results provide a general platform for strain improvement of polyol-producing microorganisms or nonconventional microorganisms in the future.