| Preparation of Knockdown Transformants of Unicellular Charophycean Alga, Closterium peracerosum-strigosum-littorale Complex
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Author:
Date:
2016-05-20
[Abstract] To prepare the knockdown transformants of the Closterium peracerosum-strigosum-littorale (C. psl.) complex, particle bombardment was applied with a newly constructed vector (pSA0104) with an endogenous constitutive promoter fused to a DNA fragment corresponding to an antisense strand of a target gene. Using a hygromycin resistance gene (aph7”), hygromycin-resistant colonies were selected. After the second screening, integration of the vector into the genome was checked by PCR and the knockdown effect was evaluated by Western blotting using a specific antibody against the target protein.
[摘要] 为了制备鬼臼藓藻（Clonsterium peracerosum-strigosum-littorale）复合物的敲低转化体，用新构建的载体（pSA0104）应用粒子轰击，所述载体具有内源性 组成型启动子，其与对应于靶基因的反义链的DNA片段融合。 使用潮霉素抗性基因（ aph7"）选择潮霉素抗性菌落。 在第二次筛选后，通过PCR检查载体到基因组中的整合，并使用针对靶蛋白的特异性抗体通过蛋白质印迹评估敲低效应。
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| Observation of Chloroplast Movement in Vallisneria
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Author:
Date:
2015-11-05
[Abstract] Chloroplasts accumulate to weak light and escape from strong light. These light-induced responses have been known from the 19th century (Böhm, 1856). Up to now, many scientists have developed different methods to investigate these dynamic phenomena in a variety of plant species including the model plant Arabidopsis thaliana, a terrestrial dicot (Wada, 2013). Especially, a serial recording to trace the position of individual chloroplast for the analysis of its mode of movement is critical to understand the underlying mechanism. An aquatic monocot Vallisneria (Alismatales: Hydrocharitaceae, Figure 1A) has contributed over a century to such investigation (Senn, 1908; Zurzycki, 1955; Seitz, 1967), because Vallisneria leaves have rectangular parallelepiped-shaped ...
[摘要] 叶绿体累积到弱光并从强光中逃逸。这些光诱发的反应从19世纪就已知（Böhm，1856）。到目前为止，许多科学家已经开发了不同的方法来研究各种植物物种中的这些动态现象，包括模拟植物拟南芥（Arabidopsis thaliana），一种陆生双子叶植物（Wada，2013）。特别是，跟踪单个叶绿体的位置，用于分析其运动模式的连续记录对于理解底层机制至关重要。水生单子叶植物Vallisneria（Alismatales：Hydrocharitaceae，图1A）在一个世纪以来已经对这种研究作出贡献（Senn，1908; Zurzycki，1955; Seitz，1967），因为Vallisneria 叶具有在单层中有序排列的长方形表皮细胞（图1B），为显微镜研究提供了优良的实验系统。在这里我们描述了一个协议为最新的时间推移成像程序来分析Vallisneria 叶绿体运动。使用这个和原型程序，相关的光感受器系统（Izutani等人，1990; Dong等人，1995; Sakai等人， ，2015），与肌动蛋白细胞骨架的关联（Dong等人，1996; Dong等人，1998; Sakai和Takagi 2005; Sakurai等人，/sh>，2005），并且已经对Ca 2 + （Sakai等人，2015）的调节作用进行了深入研究。
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