Novel plant transformation system by gene-coated gold particle introduction into specific cell using ArF excimer laser
Novel plant transformation system by gene-coated gold particle introduction into specific cell using ArF excimer laser
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DOI：
10.5511/plantbiotechnology.24.315
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发表时间：
2007-06-01
影响因子：
1.6
通讯作者：
Kobayashi, Akio
中科院分区：
文献类型：
--
作者：
Kajiyama, Shin'ichiro;Inoue, Fumihide;Kobayashi, Akio
将外源基因导入特定细胞的新方法对于特定细胞的功能分析是必需的。我们报告了一种新的完整的植物细胞的转染方法，使用ArF准分子激光诱导的冲击波引入外源基因包被的金颗粒通过细胞壁进入植物细胞。将外源基因（sGFP）包被的金颗粒随机分散在蓝猪耳组织样品的表面上;使用中空光纤和石英聚光尖端向金颗粒照射激光，以将颗粒引入靶细胞中。然后将处理过的植物组织样品在选择培养基上培养几周，并通过荧光显微镜和PCR评估转化。辐射对细胞和包被DNA的损伤不明显，转化效率与基于粒子数的基因枪转化相似。虽然这种方法需要合适的激光装置并且耗时，但它可以应用于特定靶细胞的转化实验。此外，该方法不仅可以用于导入DNA，还可以用于导入生物活性物质，例如蛋白质调节剂或抑制剂，其通过细胞壁导入细胞是困难的。
Novel methods for foreign-gene introduction into specific cells are required for the functional analysis of specific cells. We report a new transfection method for intact plant cells that uses ArF excimer laser-induced shock waves to introduce foreign gene-coated gold particles into plant cells through the cell wall. Foreign gene (sGFP)-coated gold particles were randomly scattered on the surface of Torenia tissue samples; the laser was irradiated toward the gold particles using a hollow optical fiber and a quartz light-condensing tip to introduce the particles into the target cells. Treated plant tissue samples were then cultured for several weeks on the selection medium, and transformation was assessed by fluorescence microscopy and PCR. Irradiation-induced damage to the cells and coated DNA was insignificant, and the transformation efficiency was similar to that of biolistic transformation based on the particle number. Although this method requires a suitable laser setup and is time-consuming, it could be applied in transformation experiments on specific target cells. Moreover, this method can possibly be used to introduce not only DNA but also biologically active substances such as protein regulators or inhibitors, whose introduction into cells is difficult through the cell wall.