Liu Guoquan, Campbell Bradley C, Godwin Ian D
School of Agriculture and Food Sciences, The University of Queensland, Brisbane, QLD, Australia.
Methods Mol Biol. 2014;1099:219-34. doi: 10.1007/978-1-62703-715-0_18.
Particle bombardment transformation describes the acceleration of high-velocity microparticles coated with exotic genes through the plant-protective cell walls, in order for the introduced genes to be integrated into the host genome. This technique has proven to be an effective and versatile approach towards plant genetic modification in preceding decades. Particle bombardment has been successfully applied to cereals including rice, maize, wheat, barley, and sorghum. Historically, sorghum has been considered as one of the most recalcitrant major crops with regard to successful genetic transformation; however, tremendous progress has been made in recent years. Transformation efficiency by particle bombardment has now improved from approximately 1 % to in excess of 20 % utilizing an optimized tissue culture and DNA delivery system. The protocol described in this chapter routinely generates transformants at 10-25 % efficiency within sorghum genotype Tx430. The process generally takes 11-16 weeks from initiation of immature embryos to planting of transformants. This protocol covers the operation of both the Bio-Rad PDS-1000/He System and particle inflow gun. Three factors are crucial to an efficient particle bombardment transformation system: (1) an efficient tissue culture system, (2) a highly efficient DNA delivery system, and (3) an effective selection strategy.
粒子轰击转化是指将包裹有外源基因的高速微粒加速穿过植物的保护细胞壁，以使导入的基因整合到宿主基因组中。在过去几十年里，这项技术已被证明是一种有效且通用的植物基因改造方法。粒子轰击已成功应用于包括水稻、玉米、小麦、大麦和高粱在内的谷类作物。从历史上看，高粱在成功进行遗传转化方面一直被认为是最难处理的主要作物之一；然而，近年来已取得了巨大进展。利用优化的组织培养和DNA导入系统，通过粒子轰击的转化效率现已从约1%提高到超过20%。本章所述方案在高粱基因型Tx430中常规产生效率为10%-25%的转化体。该过程通常从未成熟胚开始到转化体种植需要11-16周。本方案涵盖了Bio-Rad PDS-1000/He系统和粒子流入枪的操作。高效的粒子轰击转化系统有三个关键因素：（1）高效的组织培养系统，（2）高效的DNA导入系统，（3）有效的选择策略。
