[Low field nuclear magnetic resonance for rapid quantitation of microalgae lipid and its application in high throughput screening]
A rapid and accurate determination method of lipids in microalgae plays a significant role in an efficient breeding process for high lipid production of microalgae. Using low field nuclear magnetic resonance (LF NMR) we developed a direct quantitative method for cellular lipids in Chlorella protothecoides cells. The LF NMR signal had a linear relationship with the lipid content in the microalgae cells for both dry cell samples and algal broth samples (R2 0.99) These results indicated that we could use this method for accurate determination of microalgal lipids. Although LF NMR is a rapid and easy lipid determination method in comparison to conventional methods, low efficiency would limit its application in high throughput screening. Therefore, we developed a novel combined high throughput screening method for high lipid content mutants of C. protothecoides. Namely, we initially applied Nile red staining method for semi quantification of lipid in the pre screening process, and following with LF NMR method for accurate lipid determination in re screening process. Finally, we adopted this novel screening method in the breeding process of high lipid content heterotrophic cells of C. protothecoides. From 3 098 mutated strains 108 high lipid content strains were selected through pre screening process, and then 9 mutants with high lipid production were obtained in the re screening process. In a consequence, with heterotrophical cultivation of 168 h, the lipid concentration could reach 5 g/L, and the highest lipid content exceeded 20% (W/W) which was almost two fold to that of the wild strain. All these results demonstrated that the novel breeding process was reliable and feasible for improving the screening efficiency.
Sheng wu gong cheng xue bao Chinese journal of biotechnology