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Ca^<2+> dynamix in self-incompatibility system of plants

植物自交不亲和系统中的Ca^<2>动力

基本信息

批准号：
13640649
负责人：
IWANO Megumi
金额：
$ 2.11万
依托单位：
Nara Institute of Science and Technology
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2001
资助国家：
日本
起止时间：
2001 至 2002
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13640649/
关键词：
Arabidopsis Brassica Ca^<2+> oscillation transformation Self-incompatibility yellow cameleon 自家不和合成 Aravidopsis

项目摘要

In order to clarify the role of Ca^<2+> in pollen-stigma interaction of Brassicaceae, dynamic movement of Ca^<2+> in pollen grains and papilla cells during pollination process should be examined precisely and with time. Typically, ion movements in a cell have been analyzed by injecting ion sensitive dye into the cell, but the microinjection often causes physical damage to the cell and it is difficult to inject the dye to a pollen grain because pollen grain is covered with hard coats, such as exine layer. The fluorescent protein indicator for Ca^<2+>, "yellow cameleons" (YC) has been developed to combine the advantages of molecular biological targeting and fluorescence readout. In this study, first two kinds of YC3.1 chimeric genes containing Act1 promoter region or SLG promoter region were transformed into Arabidopsis thaliana. Then two kinds of transgenic plants, which expressed YC3.1 in pollen grains or papilla cells, respectively, were obtained. After the pollen grain was pollinated to the papilla cell using micromanipulator, Ca^<2+> dynamics were monitored with time. Before germination, Ca^<2+> accumulated at the site, which the pollen grain contacted to the papilla cell. In the germinated pollen tube, faster movement of Ca^<2+> was observed, compared with that of in vitro germinated pollen tube. On the other hand, Ca^<2+> movement in the papilla cell was slower than that in pollen grains. Next the YC3.1 chimeric genes were transformed into Brassica rapa. The transgenic plant, which expressed YC3.1 in the pollen grains, was obtained. As a result, dynamic movement of Ca^<2+> in the pollen grains was observed only in cross-pollination. Dynamic movement of Ca^<2+> in pollen grains and papilla cells during pollination process could be monitored precisely and with time using transgenic plants, which expressed YC genetically.

为了阐明Ca^<2+>在菊科植物花粉-柱头相互作用中的作用，需要对授粉过程中花粉粒和乳突细胞中Ca^<2+>的动态变化进行精确的、随时间变化的研究。通常，通过将离子敏感染料注射到细胞中来分析细胞中的离子运动，但是显微注射经常对细胞造成物理损伤，并且难以将染料注射到花粉粒中，因为花粉粒被硬的外套（例如外壁层）覆盖。Ca^<2+>的荧光蛋白指示剂“黄驼龙”（YC）已被开发以联合收割机结合分子生物学靶向和荧光读出的优点。本研究将前两种分别含有Act 1启动子区和SLG启动子区的嵌合基因YC3.1转化到拟南芥中。获得了在花粉粒和乳突细胞中分别表达YC 3.1的转基因植株。用显微操作器将花粉粒授粉到乳突细胞后，观察Ca^<2+>动态随时间的变化。萌发前，Ca^2+在花粉粒与乳突细胞接触的部位积累.与离体萌发的花粉管相比，萌发的花粉管中Ca^<2+>的移动速度更快。另一方面，乳突细胞中Ca^2+的运动比花粉粒中慢.然后将YC 3.1嵌合基因转化到芜菁中。获得了在花粉粒中表达YC 3.1的转基因植株。结果表明，花粉粒中Ca^<2+>的动态运动仅在异花授粉中观察到。利用转YC基因植株可以精确地监测授粉过程中花粉粒和乳突细胞中Ca^2+随时间的动态变化。