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用低能氩离子束介导将构建的pCAMBIA1308质粒载体上携带的几丁质酶基因(RCH8)导入3个小麦栽培品种扬麦158, 皖9210, 皖麦32号的成熟胚细胞. 来自成熟胚的初生愈伤组织首先经过含有20~30 mg/L潮霉素(hygromycin, Hm)的培养基的筛选培养, 获得的抗性愈伤组织转入Hm浓度为10~20 mg/L的培养基上进行分化培养, 3个小麦品种都获得了再生植株. 再生苗PCR和PCR-Southern检测结果显示, 绝大部分再生植株为阳性的转化植株. Southern杂交检测表明外源RCH8基因已整合到小麦基因组中. 植株转化率在不同品种间有所不同, 同一品种经不同剂量的氩离子束注入后, 转化效果也存在较大差异: 皖麦32号最高转化率可达3.8%; 扬麦158在5种离子注入剂量下的转化率为0.5%~2.5%; 皖9210的转化率为0.5%~1.4%. 对转化二代植株中几丁质酶基因的表达情况的检测结果显示, 有两个转化株系的幼苗叶片细胞粗提液对两种小麦赤霉病菌毒菌株表现出抗性.
The chitinase gene (RCH8) carried on the constructed pCAMBIA1308 plasmid vector was introduced into mature wheat embryos of three wheat cultivars Yangmai 158, Wan 9210 and Wanmai 32 by using low energy argon ion beam. Primary callus was first cultured in a medium containing 20 ~ 30 mg / L hygromycin (Hm), and the resistant calli were transferred to medium with Hm concentration of 10 ~ 20 mg / L The regenerated plants were obtained by PCR and PCR-Southern analysis.The results showed that most of the regenerated plants were positive transformed plants.Experimental results of Southern hybridization showed that the exogenous RCH8 gene was integrated into the genome of wheat . The plant transformation rate was different among different cultivars. The same variety had different transformation effects after different doses of argon ion beam injection: the highest conversion of Wanmai 32 was 3.8% The conversion rate was 0.5% ~ 2.5% at ion implantation dose and 0.5% ~ 1.4% at Anhui 9210. The results of chitinase gene expression in the second generation of transformed plants showed that there were two transformants Department of seedling leaf cell crude extract pairs Two strains of Fusarium graminearum showed resistance.