农杆菌介导创伤胚转化的应用

材料，农杆菌菌株LBA4404（含质粒P3301ubiAc）侵染转化燕麦、玉米和高粱的萌发种子，优化转化条件，探索燕麦、玉米和高粱遗传转化体系，探讨菌液浓度、种子萌发时间、超声波功率、乙酰丁香酮和草丁膦浓度对3种作物的影响并进行比较分析。[结果]农杆菌菌液浓度为0.6>OD600≥0.4 时转化效果最佳;第1次超声波功率为900 W，处理时间为10 min;第2次超声波功率为200 W，处理时间为4 min效果最佳。转化效率最高;添加乙酰丁香酮对燕麦、高粱和玉米的发芽有明显促进作用，但转化效果有明显区别;燕麦、玉米和高粱的转化植株对草丁膦的耐受性差异不大，与非转化植株相比，0.8 mg/L 除草剂胁迫下燕麦、玉米和高粱种子萌发受到一定程度的抑制。用1 mg/L 除草剂溶液对T₀和T₁代转化植株幼苗进行筛选，转化植株的存活率高于对照。[结论]经除草剂筛选和PCR对T₀和T₁代幼苗检测，初步证明获得了转化植株。

关键词 植物转化;燕麦;玉米;高粱

中图分类号 S188  文献标识码 A  文章编号 0517-6611（2015）03-025-04

Trauma Embryo of Genetic Transformation System with Agrobacterium Mediated Method

ZHANG Lijun1，2， LIU Longlong¹，ZHANG Jianzhen²， CUI Lin1* et al

（1. Crop germplasm resources research institute， Shanxi Academy of Agricultural Sciences; Key Laboratory of Crop Gene Resources and Germplasm Enhancement on Loess Plateau， Ministry of Agriculture， P. R. China， Taiyuan， Shanxi  030031;2. Research Institute of Applied Biology，Shanxi University，Taiyuan，Shanxi  030001）

Key words [Objective]The purpose was to optimize agrobacteriummediated plant germination seed transformation method，increase conversion rate， breed GM varieties with specific functions， and improve viability and adaptability of crops. [Method]This paper consulted agrobacteriummediated plant germination seed transformation method and selected germinating seeds of oat， maize and sorghum as test materials. Germinating seeds of oat， maize and sorghum were infected and transformed by agrobacterium strain， which included plasmid P3301ubiAc. Transformation conditions were optimized. Genetic transformation systems of oat， maize and sorghum were studied. Concentration of bacterium， seed germinating time， ultrasonic power， and influence of concentration of acetosyringone and glufosinate on the above three crops were compared and analyzed. [Result]The experiment showed that when 0.6>OD600≥0.4， transformation achieved the best effect; Ultrasonic power was 900 W at the first time and treatment time was 10 minutes; Ultrasonic power was 200 W at the second time and transformation achieved the best effect when treatment time was 4 minutes; Acetosyringone could affect germination of oat， sorghum and maize significantly. It could increase transformation rate of maize and lower that of sorghum， but for oat， its effect was not obvious; in terms of tolerance to glufosinate， the difference was not significant among transformed plants of oat， maize and sorghum. Compared with nontransformed plants， seed germination of oat， maize and sorghum were restrained to some degree under the stress of 0.8 mg/L herbicide solution. With 1 mg/L herbicide solution to select seedlings of T₀ and T₁ transformed plants， the result showed survival rates of transformed plants is higher than those of control group. [Conclusion ]After herbicide selection and PCR test on T₀ and T₁ seedlings， it was proved preliminarily that transformed plants were got.

推荐访问： 杆菌 创伤 转化

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