Establishment of a Genetic Transformation System for the Marine Pennate Diatom Fistulifera sp Strain JPCC DA0580-A High Triglyceride Producer
Establishment of a Genetic Transformation System for the Marine Pennate Diatom Fistulifera sp Strain JPCC DA0580-A High Triglyceride Producer
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DOI：
10.1007/s10126-012-9457-0
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发表时间：
2013-02-01
影响因子：
3
通讯作者：
Tanaka, Tsuyoshi
中科院分区：
文献类型：
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作者：
Muto, Masaki;Fukuda, Yorikane;Tanaka, Tsuyoshi
采用基因枪法建立了海洋羽纹硅藻Fistulifera sp. JPCC DA 0580的遗传转化体系。菌株JPCC DA 0580最近已被鉴定为从1，393株海洋微藻的培养物收集物中最高的甘油三酯（60%w/w）生产者，并且预期其是生物柴油燃料的可行来源。使用绿色荧光蛋白基因（gfp）和编码新霉素磷酸转移酶II（npt II）的基因优化菌株JPCC DA 0580的转化条件。当钨颗粒（直径0.6 μ m）用于微粒轰击时，获得最有效的转化率。研究了内源性和外源性启动子对npt II表达的影响。与外源启动子相比，内源启动子更有效地获得转化子。Southern杂交分析表明npt Ⅱ已整合到核基因组中。这种遗传操作技术应该使我们能够了解这种菌株中高甘油三酯积累的机制，从而有助于提高BDF的生产。
A genetic transformation system for the marine pennate diatom, Fistulifera sp. JPCC DA0580, was established using microparticle bombardment methods. Strain JPCC DA0580 has been recently identified as the highest triglyceride (60 % w/w) producer from a culture collection of 1,393 strains of marine microalgae, and it is expected to be a feasible source of biodiesel fuel. The transformation conditions for strain JPCC DA0580 were optimised using the green fluorescent protein gene (gfp) and the gene encoding neomycin phosphotransferase II (nptII). The most efficient rate of transformation was attained when tungsten particles (0.6 mu m in diameter) were used for microparticle bombardment. The effect of endogenous and exogenous promoters on the expression of nptII was examined. Endogenous promoters were more efficient for obtaining transformants compared with exogenous promoters. Southern hybridisation analysis suggested that nptII integrated into the nuclear genome. This genetic manipulation technique should allow us to understand the mechanisms of high triglyceride accumulation in this strain, thereby contributing to improving BDF production.