High-throughput Screening For High-yield Strains Of Monascus Pigment

High-throughput screening platform inculdes two complementary technical systems: high-throughput culture technology and high-throughput screening technology. An economical and rapid high-throughput primary screening strategy was developed for high-aerobic Monascus purpureus cultivation. The problem of mycelium clustering in 24-well plates was solved by using mycelium suspension as the culture seed. The culture volume and evaporation characteristics of the 24-well plates was also investigated. The good correlation of fermentation results between differing-scale cultivations confirmed the feasibility of utilizing the 24-deep microtiter plates (MTPs) as a scale-down tool instead of shake flasks for culturing this Monascus pigment producing microbes. Supernatants obtained by a rapid centrifugation step after fermentation broth was extracted with ethanol, then it was evaluated by 96 deep-well and microplate reader at 530 nm, which is more rapid and sensitive. The whole technique system is simpler and more stable.To operated and verified the established high throughput screening platform, Monascus purpureus was induced multiplex mutation with UV-LiCl for screening a mutant with high yield of pigment. Nine high mixture strains in the plate wells were obtained after compound mutagenesis using the high throughput screening method. The traditional dilution-plate method was employed to further isolate the high-yield single colony from the selected high-yield mixture. The pigment yield of D39-4 increased by 51.5%, and had stable inheritance after verification in the shake flask experiments.In order to increase the production of the Monascus purpureus D39-4, the fermentation medium was optimized using response surface methodology (RSM). Formula of the culture medium was as follow:Glucose 51.42 g/L, peptone 4.91 g/L, NaNO3 1.00 g/L, KH2PO4 1.00 g/L, MgSO4·7H2O 1.00 g/L. The final yields of Shake flasks pigment was 265.8 U/ml, increasing 30% than that in the medium optimized before.