Protocol: a medium-throughput method for determination of cellulose content from single stem pieces of Arabidopsis thaliana.
Protocol: a medium-throughput method for determination of cellulose content from single stem pieces of Arabidopsis thaliana.
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DOI：
10.1186/s13007-015-0090-6
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发表时间：
2015
期刊：
影响因子：
5.1
通讯作者：
Turner S
中科院分区：
文献类型：
--
作者：
Kumar M;Turner S
木质纤维素生物质是重要的可再生资源，可用于生物燃料和材料。植物如何合成纤维素还不完全清楚。已知在质膜中移动的纤维素合成酶复合体（CSC）合成纤维素。CESA蛋白是CSC的核心成分。在拟南芥中，蛋白质的体外诱变，然后对缺乏该基因的突变体进行互补分析，是研究任何生物过程（包括纤维素生物合成）的重要工具。对大量植物的分析对于这些类型的研究至关重要。通过在方案的各个阶段使用抽吸而不是离心来去除液体，我们能够增加该方法的通量并最小化样品损失。作为一个测试案例，我们确定了纤维素含量的野生型和次生壁cesa突变体横跨长度的初级拍摄，这是喜欢在7周龄的植物是相当均匀的。此外，我们发现，单突变体的纤维素含量是可比的高阶突变体。在这里，我们描述了一个中等吞吐量的适应Updegraff的方法，使我们能够确定每周200个样品的纤维素含量。
Lignocellulosic biomass is an important renewable resource for biofuels and materials. How plants synthesise cellulose is not completely understood. It is known that cellulose synthase complex (CSCs) moving in the plasma membrane synthesise the cellulose. CESA proteins are the core components of CSC. In Arabidopsis, in vitro mutagenesis of proteins followed by complementation analysis of mutants lacking the gene represents an important tool for studying any biological process, including cellulose biosynthesis. Analysis of a large number of plants is crucial for these types of studies. By using aspiration rather than centrifugation to remove liquids during various stages of protocol, we were able to increase the throughput of the method as well as minimise the sample loss. As a test case, we determined cellulose content of wild type and secondary wall cesa mutants across the length of primary shoot which was fond to be rather uniform in 7-week-old plants. Additionally, we found that the cellulose content of single mutants was comparable to the higher order mutants. Here we describe a medium-throughput adaptation of Updegraff’s method that allowed us to determine cellulose content of 200 samples each week.