影响基因枪转化小麦的几个因素

FACTORS AFFECTING WHEAT TRANSFORMATION EFFICIENCY BY PARTICLE BOMBARDMENT

对影响基因枪转化小麦的主要参数进行优化 ,结果表明 ,包裹子弹的沉淀剂用PEG40 0 0和Ca(NO3 ) 2 分别代替亚精胺和CaCl2 ,可显著地提高GUS基因的瞬时表达 .幼穗的最佳预培养时间为 1d ,轰击距离为 9cm ,而幼胚分别为 14d和 12cm .对预培养 14d的幼胚 ,采用 0 2 5 μgDNA和 12 5

The major parameters controlling DNA delivery to wheat (Triticum aestivum. L) by particle bombardment were investigated, with four wheat cultivars from Southwest area of China as target tissues. The results indicated that when spermidine and CaCl 2,the precipitants for coating particles, were replaced with PEG4000 and Ca (NO 3) 2 respectively, the frequency of GUS transient expression was improved significantly. The bombardment parameters for immature spikes and immature embryos were tuned, and it was found that the transformation frequency was relatively high when immature spikes pre cultured in vitro for 1 day was used as target tissues with the bombardment distance in 9cm, and immature embryos for 14 days with the bombardment distance in 12cm. In addition, the dosage of gold powder and DNA per bombardment was one of the key factors for the successful transformation. In this study, using 14 days pre cultured immature embryos as target tissues, 125μg gold powder coated by 0.5μg DNA was the best combination.