Study on Male Sterility of Plants by Direct Transfer of Mitochondria and mitochondrial DNA

粒子轰击已经被先前的作者(例如，Svab等人)证明是一种有效的细胞器转化方法。《美国国家科学院院刊》87：，8526-8530,1990年)。在这项研究中，我们研制了自己的粒子枪装置，并尝试转化高等植物的线粒体。我们还没有成功地完成线粒体的转化。然而，我们得到了一些基本的结果，这些结果将对未来细胞器转化的研究有用。我们已经开发了由氮气压力和压缩空气压力驱动的粒子枪装置。氮气压力驱动装置的最大加速压力(即用于加速塑料弹丸的压力)为25 kg/cm2，而气动粒子炮装置的最大加速压力超过220 kg/cm2。使用气动粒子炮，弹丸的最大速度超过440m/s2。利用这些设备，我们已经证明了质粒DNA的成功导入，以及导入的外源基因(如GUS基因)可以在烟草、大豆和拟南芥(双子叶)以及水稻和小麦(单子叶)等植物的完整组织和培养细胞中成功表达。利用这两种粒子枪装置，成功地实现了NPTII基因烟草培养细胞的稳定转化。我们还将NPTII基因导入拟南芥的根细胞中，并选择了卡那霉素抗性的植株。在卡那霉素和结实种子的存在下，所选的芽成功地发育成完整的植株(到目前为止已经获得了20多株这样的植株)。

Particle bombardment has been shown to be a powerful method for the transformation of organelles by the previous authors (e. g., Svab et al. PNAS 87 : , 8526-8530, 1990). In this study we developed our own particle gun device and attempted to transform mitochondria of higher plants. We have not yet succeeded in transformation of mitodondria. However, we obtained some fundamental results that will be useful for studies on organeLle transformation in future.1. We have developed particle gun devices that are driven by nitrogen gas pressure and compressed air pressure. Nitrogen-gas-pressure-driven device had a maximum accelerating pressure (i. e. pressure that was used to accelerate plastic projectiles) of 25 kg/cm^2, while pneumatic particle gun device had a maximum accelerating pressure of more than 220 kg/cm^2. With the pneumatic particle gun, the maximum velocity of projectile was more than 440 m/s.2. Using these devices, we have shown that plasmid DNA can successfully be introduced and the introduced foreign genes (e. g. GUSgene) can successfully be expressed in intact tissues as well ascultured cells of various plants such as tobacco, soybean and Arabidopsis (dicots) and rice and wheat (monocots).3. We succeeded in stable transformation of cultured cells of tobacco with NPTII gene by use of these two particle gun devices. We also delivered the NPTII gene into root cells of Arabidopsis and selected kanamycin-resistant shoots. The selected shoots successfully developed into complete plants (more than 20 such plants have so far been obtained) in the presence of kanamycin and set seeds.