Optimization of a simple, accurate and low cost method for starch quantification in green microalgae
Optimization of a simple, accurate and low cost method for starch quantification in green microalgae
复制标题
优化简单、准确、低成本的绿色微藻淀粉定量方法
作者：
T. Yong;Chia;Ching
脂类和淀粉是生物能源生产的重要原料。绿色微藻中脂类和淀粉生物合成的遗传研究近年来引起了人们的广泛关注。在这些研究中，需要对脂质和淀粉进行定量，以阐明因果关系。虽然全世界都用类似的方法测定脂质，但绿色微藻中的淀粉已经使用各种方法测量，这些方法在准确性上存在缺陷或成本高。建立了一种简便、准确、低成本的绿色微藻淀粉定量分析方法。该程序包括细胞的快速冷冻、色素的溶剂提取、134 °C高压灭菌和淀粉的葡糖淀粉酶双消化，然后使用二硝基水杨酸试剂进行葡萄糖测定。优化该程序以定量小体积绿色微藻培养物中的淀粉。使用该程序进行淀粉定量的准确度为102.3 ± 2.5%（平均值± SD，n = 6），如使用玉米淀粉作为内部对照所示。工作协议可在http://dx.doi.org/10.17504/protocols.io.2mhgc36上获得。该方法克服了目前绿色微藻淀粉定量方法存在的不准确、成本高等问题。这种方法将提供一个机会，比较遗传，生理或培养操作对淀粉生产力的绿色微藻在不同的实验室中阐明，这是必不可少的在提高微藻中的脂质生产力的研究。
Lipids and starch are important feedstocks for bioenergy production. Genetic studies on the biosyntheses of lipids and starch in green microalgae have drawn significant attention recently. In these studies, quantifications of lipids and starch are required to clarify the causal effects. While lipids are assayed with similar procedures worldwide, starch in green microalgae has been measured using various methods with deficiencies in accuracy or high cost. A simple, accurate and low cost procedure for routine quantification of starch in green microalgae was developed. This procedure consists of quick-freezing of the cells, solvent extraction of the pigments, 134 °C autoclaving and glucoamylase double digestions of starch, followed by a glucose assay using the dinitrosalicylic acid reagent. This procedure was optimized to quantify starch in small volumes of green microalgal culture. The accuracy of starch quantification using this procedure was 102.3 ± 2.5% (mean ± SD, n = 6), as indicated by using cornstarch as internal controls. The working protocol is available at http://dx.doi.org/10.17504/protocols.io.2mhgc36. This quantification approach overcomes the current problems in the starch quantification of green microalgae such as inaccuracy and high cost. This approach would provide an opportunity to compare the effects of genetic, physiological or cultivation manipulations on the productivity of starch in green microalgae elucidated in different labs, which is essential in the enhancement of lipid productivity studies in microalgae.