Armaleo D, Ye G N, Klein T M, Shark K B, Sanford J C, Johnston S A
Department of Botany, Duke University, Durham, NC 27706.
Curr Genet. 1990 Feb;17(2):97-103. doi: 10.1007/BF00312852.
Tungsten microprojectiles coated with nucleic acid and accelerated to velocities of approximately 500 m/s, can penetrate living cells and tissues with consequent expression of the introduced genes (Klein et al. 1987). Saccharomyces cerevisiae is used here as a model system to define the basic parameters governing the biolistic (biological-ballistic) delivery of DNA into cells. Among the physical factors affecting the efficiency of the process in yeast are the microprojectile's constitution, size, concentration and amount, and the procedure used for binding DNA to it. The biological parameters that affect the process include the cell's genotype, growth phase, plating density, and the osmotic composition of the medium during bombardment. By optimizing these physical and biological parameters, rates of transformation between 10(-5) and 10(-4) were achieved. Stable nuclear transformants result primarily from penetration of single particles of 0.5-0.65 micron in diameter, delivering on average 10-30 biologically active plasmids into the cell. The tungsten particles detectably increase the buoyant density of the transformants' progenitors.
涂有核酸并加速至约500米/秒速度的钨微射弹，能够穿透活细胞和组织，从而使导入的基因得以表达（克莱因等人，1987年）。这里使用酿酒酵母作为模型系统，以确定将DNA通过生物弹道法导入细胞的基本参数。影响酵母中该过程效率的物理因素包括微射弹的组成、大小、浓度和数量，以及将DNA与之结合所采用的程序。影响该过程的生物学参数包括细胞的基因型、生长阶段、平板接种密度以及轰击期间培养基的渗透成分。通过优化这些物理和生物学参数，转化效率达到了10⁻⁵至10⁻⁴之间。稳定的核转化体主要源于直径为0.5 - 0.65微米的单个粒子的穿透，平均将10 - 30个具有生物活性的质粒导入细胞。钨粒子可显著提高转化体祖细胞的浮力密度。
