兼养培养对三种典型微藻生长与胞内组分及脂质合成相关基因表达的影响研究
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摘要
微藻生长迅速、生物量高并且富含油脂,因此是生产生物柴油的理想原料。许多微藻在兼养培养条件下与光能自养条件下相比具有更高的生长速率。在本文中对三种微藻眼点拟微绿球藻(Nannochloropsis oculata HQW03)、盐生杜氏藻(Dunaliella salina HQW04)和小球藻(Chlorella sorokiniana CS-01)在以葡萄糖为有机碳源的兼养培养条件下的生长、脂类和蛋白质含量的变化进行了研究。结果表明当培养基中葡萄糖浓度达到最适浓度时,眼点拟微绿球藻(Nannochloropsis oculata HQW03)、盐生杜氏藻(Dunaliella salina HQW04)和小球藻(Chlorella sorokiniana CS-01)的生物量分别是自养培养条件下的1.4、2.2和4.2倍。同时,脂类和蛋白质含量也有所增加(眼点拟微绿球藻：23%和201%；盐生杜氏藻：20%和43%；小球藻：86%和215%)。通过比较三种微藻脂类产量,平均每天的脂类产量以及添加葡萄糖后增加的脂类产量发现小球藻CS-01与其他两种微藻相比在许多方面都是最理想的藻种,如生长速率,脂类产量以及生产速率和葡萄糖利用率等。
通过Real-time PCR对小球藻CS-01在不同浓度葡萄糖培养条件下的三种脂类合成相关基因(accD, accl和rbcL)的表达差异进行了研究。结果表明：异质型乙酰辅酶A羧化酶是小球藻CS-01脂类合成的关键酶,accD基因的表达水平能够反映微藻在兼养条件下稳定期脂类含量的情况；另外,编码同质型乙酰辅酶A羧化酶的accl基因的表达水平很低甚至不表达；rbcL基因的不同表达水平表明小球藻CS-01在兼养培养条件下,光能自养和化能异养生长不是独立进行的,并且有机碳源的利用可以减小光合作用效率。
Microalgae can grow extremely rapidly and are rich in oils that are a suitable feedstock for biodiesel production. Many microalgae exhibit higher growth rates under mixotrophic conditions than under photoautotrophic conditions. In the present study, three species of microalgae Nannochloropsis sp.. HQW03, Dunaliella salina HQW04 and Chlorella sorokiniana CS-01 under mixotrophic culture with glucose were compared with photoautotrophic culture in terms of growth, lipid and protein contents, The results show that with optimal concentrations of glucose for growth in the media, the biomass productivity of Nannochloropsis sp.., Dunaliella salina and Chlorella sorokiniana were found to be 1.4,2.2 and 4.2 fold of that obtained under photoautotrophic conditions; meanwhile, the contents of lipid and protein were increased (Nannochloropsis sp..:23% and 201%; Dunaliella salina:20% and 43%; Chlorella sorokiniana:86% and 215%, respectively). Upon comparison of the lipid yield, average lipid yield per day, and additional lipid yield with glucose of the three microalgae, Chlorella sorokiniana CS-01 proved to be the best organism for lipid production in many aspects, such as growth rate, lipid yield and productive rate, and glucose utilization efficiency.
Furthermore, the expression level of accD, accl, rbcL genes in Chlorella sorokiniana CS-01 with various glucose concentrations were also studied by real-time PCR. The results show:heteromeric ACCase is one of key enzymes for lipid synthesis in Chlorella sorokiniana, as the expression level of accD can reflect lipid contents of algae in stationary phase of mixotrophic growth; Additionally, the acc1 gene coding homomeric ACCase remains constitutively expressed at low levels or even not expressed; The differential expression of the rbcL gene suggests that photoautotrophic and heterotrophic growth of Chlorella sorokiniana CS-01 cannot be proceed independently under mixotrophic conditions, and utilization of an organic carbon source will reduce the photosynthesis efficieny.
通过Real-time PCR对小球藻CS-01在不同浓度葡萄糖培养条件下的三种脂类合成相关基因(accD, accl和rbcL)的表达差异进行了研究。结果表明：异质型乙酰辅酶A羧化酶是小球藻CS-01脂类合成的关键酶,accD基因的表达水平能够反映微藻在兼养条件下稳定期脂类含量的情况；另外,编码同质型乙酰辅酶A羧化酶的accl基因的表达水平很低甚至不表达；rbcL基因的不同表达水平表明小球藻CS-01在兼养培养条件下,光能自养和化能异养生长不是独立进行的,并且有机碳源的利用可以减小光合作用效率。
Microalgae can grow extremely rapidly and are rich in oils that are a suitable feedstock for biodiesel production. Many microalgae exhibit higher growth rates under mixotrophic conditions than under photoautotrophic conditions. In the present study, three species of microalgae Nannochloropsis sp.. HQW03, Dunaliella salina HQW04 and Chlorella sorokiniana CS-01 under mixotrophic culture with glucose were compared with photoautotrophic culture in terms of growth, lipid and protein contents, The results show that with optimal concentrations of glucose for growth in the media, the biomass productivity of Nannochloropsis sp.., Dunaliella salina and Chlorella sorokiniana were found to be 1.4,2.2 and 4.2 fold of that obtained under photoautotrophic conditions; meanwhile, the contents of lipid and protein were increased (Nannochloropsis sp..:23% and 201%; Dunaliella salina:20% and 43%; Chlorella sorokiniana:86% and 215%, respectively). Upon comparison of the lipid yield, average lipid yield per day, and additional lipid yield with glucose of the three microalgae, Chlorella sorokiniana CS-01 proved to be the best organism for lipid production in many aspects, such as growth rate, lipid yield and productive rate, and glucose utilization efficiency.
Furthermore, the expression level of accD, accl, rbcL genes in Chlorella sorokiniana CS-01 with various glucose concentrations were also studied by real-time PCR. The results show:heteromeric ACCase is one of key enzymes for lipid synthesis in Chlorella sorokiniana, as the expression level of accD can reflect lipid contents of algae in stationary phase of mixotrophic growth; Additionally, the acc1 gene coding homomeric ACCase remains constitutively expressed at low levels or even not expressed; The differential expression of the rbcL gene suggests that photoautotrophic and heterotrophic growth of Chlorella sorokiniana CS-01 cannot be proceed independently under mixotrophic conditions, and utilization of an organic carbon source will reduce the photosynthesis efficieny.
引文
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[3]Khan S.A., Rashmi, Hussain M.Z., et al. Prospects of biodiesel production from microalgae in India[J]. Renewable and Sustainable Energy Reviews,2009, 13:2361-2372.
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[5]朱建芳,钱伯章.生物柴油生产现状及技术进展[J].天然气与石油,2007,25(3)：49-52.
[6]苏敏光,于少明,吴克,等.生物柴油制备方法及其质量标准现状[J].包装与食品机械,2008,26(3)：20-25.
[7]宋东辉,侯李君,施定基.生物柴油原料资源高油脂微藻的开发利用[J].生物工程学报,2008,24(3)：341-348.
[8]夏金兰,万民熙,王润民,等.微藻生物柴油的现状与进展[J].中国生物工程杂志,2009,29(7)：118-126.
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[17]魏东,张学成,隋正红,等.氮源和N/P对眼点拟微球藻的生长、总脂含量和脂肪酸组成的影响[J].海洋科学,2000,24(7)：46-51.
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