小球藻的甲基磺酸乙酯诱变及产EPA的条件研究

小球藻是海水养殖系统中常用的单细胞微藻,繁殖能力强,易于规模化培养,且可合成不饱和脂肪酸EPA、DHA等多种活性物质,在医疗和保健品开发中具有很高的应用价值。目前商业化培养的藻种多从自然界中直接获得,活性物质的产量较低且藻种易退化。为了获得EPA产量更高的藻种,该研究利用0.6%EMS对小球藻进行诱变,利用尼罗红染色法进行初筛并通过单细胞分离技术得到1株突变株EC1,通过气象色谱测定其EPA产量。结果表明:与出发藻株相比,突变株EPA(二十碳五烯酸)产量提高了8.97%。根据单因素试验确定突变株生长及产EPA的合适培养条件,再通过正交试验筛选出培养条件的优化组合,表明突变藻EC1株产EPA的较适条件为NaNO_375 mg·L~-1,p H7.5,昼夜温度17~15℃,接种量为12%,在此条件下培养7d其EPA的产量可达25.38 mg·g~-1,传代实验表明突变藻株具有较好的遗传稳定性。该研究结果为进一步利用小球藻规模化生产EPA奠定了基础。

EMS mutagenesis of Chlorella and the conditions of producing EPA research

Chlorella vulgaris is a single-cell microalgae and most frequently used in aquaculture because it is easy for large-scale cultivation and could generate PUFAs such as EPA and DHA, which possess high value in medical and health care products. But the microalgae for commercial training were directly obtained from the nature, whose active material production was low and easy to degradate. In order to obtain algal strains of high EPA production, C. vulgaris was mutated by 0.6%EMS. Using Nile red staining for preliminary screening and then selected by single-cell clone meth-od, the mutant strain EC1 was isolated from 200 mutant clones. The EPA yield of EC1 was measured by gas chromatog-raphy. The yield increased by 8.97% compared with the parent strain. The appropriate condition for the mutant strain culturing and its EPA production was determined by single factor experiment. Then the optimize condition assembly was determined by orthogonal test. The most suitable culture conditions for EPA production of EC1 included NaNO3 75 mg?L-1, pH7.5, day-night temperature 17-15 ℃, 12% inoculation quantity and training for 7 d. The EPA yield of EC1 could reach 25.38 mg?g-1 under the suitable culture condition. The results of subculture test showed that the mutant strain possessed good hereditary stability. The EPA yield showed no significant difference from the first generation to the sixth generation. The research would lay the foundation for the utilization of the C. vulgaris.