摘要
[目的]以破囊壶菌Thraustochytrium sp.FJN-10为研究对象,研究不同培养温度和变温条件对菌体生物量、油脂含量、脂肪酸组分、关键基因转录以及蛋白质组的影响。[方法]通过摇瓶实验测定干重研究菌株的生长,提取油脂并经液相色谱分析脂肪酸组分;荧光定量PCR和二维电泳研究脂肪酸合成途径关键的碳链延长酶、脱饱和酶的转录水平和蛋白质的差异表达。[结果]28℃培养时,生物量达13.6 g/L,DHA占总脂肪酸含量达32.41%;15℃培养时,生物量为9.8 g/L,DHA占总脂肪酸含量达56.08%。变温培养下生物量最高可达13.9 g/L、油脂含量及DHA含量在较高的水平。28℃降至15℃时,△4脱饱和酶和△6延长酶基因基因的转录分别提高了3.9和2.5倍。[结论]变温条件下菌株生物量在11.8~13.6 g/L,菌体稳定期延长,DHA的含量可达45%以上。可为今后DHA的产业化提供基础数据。
[ Objective] The study object is marine microorganism Thraustochytrium sp. FJN - 10. Attempts were made to anal- ysis the cell growth,lipid content and fatty acid profile of Thraustochytrium sp. FJN - 10 at different temperatures and tempera- ture shift. [ Methods] Dry weight method was used to measure biomass determination of Thraustochytrium sp. FJN - 10. The fat- ty acid profiles of strain were analyzed by liquid chromatogram. Real - time quantitative PCR was used to analyse gene tran- scription level of key enzymes in fatty acids synthetic pathway. Proteome quantitative changes of Thraustochytrium sp. FJN - 10 at different temperatures was analysed by 2 - D electrophoresis. [ Results] The decrease of temperature from 28~C to 15~C sup- pressed cell growth but enhanced the production of polyunsaturated fatty acids （PUFAs） ,which lead to an increase of the yield of DHA from 32.41% up to 56.08%. In addition,temperature shift under optimum conditions further enhanced the growth and the production of PUFAs. Gene expression level of a4desaturase, A5 elongase, A5 desaturase and A6 elongase were elucida- ted by real - time PCR. The levels of A4 desaturase and A6 elongase mRNA expression were significantly increased in low culture temperature. [ Conclusion] The strain had great biomass about 11.8 - 13.6 g/L and high content of DHA about 45% in PUFAs when temperature shift. These results above will also be helpful to study biochemical characteristics and be useful for further production of DHA of this strain.
出处
《生物技术》
CAS
CSCD
北大核心
2015年第4期384-390,共7页
Biotechnology
基金
国家自然科学基金项目("海洋破囊壶菌二十二碳六烯酸生物合成相关碳链延长酶基因的克隆与表达"
No.30370028)
福建省自然科学基金重大项目("发酵法生产二十二碳六烯酸的关键技术开发及产业化"
No.2011H6007)资助
关键词
破囊壶菌
不饱和脂肪酸
二十二碳六烯酸
温度
荧光定量PCR
二维电泳
Thraustochytrium sp. FJN - 10, polyunsaturated fatty acids, docosabexaenoicacid, temperature, Real - time quanti- tative PCR,2 - D electrophoresis
