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目的:将aFGF与红花的油体蛋白融合,转入红花中,直接通过提取红花油来实现红花与aFGF的双重功效。方法:以不同日龄的红花子叶为外植体,采取正交试验设计的方法,摸索出红花再生体系的最佳条件。然后通过农杆菌介导法将aFGF基因转入到红花中,对不同浓度的抗生素进行筛选,进行PCR鉴定,计算阳性率。结果:根据不同的激素配比,确定了红花分化的最佳培养基为MS+BA1.0 mg.L-1+NAA0.2 mg.L-1;生根的最适培养基为1/4 MS+NAA2.0 mg.L-1+IAA0.5 mg.L-1。3种不同种类和浓度的抑菌抗生素对红花浸染后的抑菌效果略有不同,结合抑菌效果,污染率,芽的分化率以及成本问题等因素的综合考虑,抑菌抗生素选择用Tim,质量浓度为400 mg.L-1即可抑制细菌的生长,同时有利于芽的分化;hyg的筛选压力则选择6 mg.L-1为它的临界浓度;对8棵转化植株进行鉴定,阳性率为25%。结论:通过器官发生途径,确定出红花分化与生根的最佳激素组合与配比,建立了红花的再生体系;确定出最适的抑菌抗生素与筛选抗生素的浓度;经过筛选获得的再生植株,通过PCR检测,部分扩增出aFGF目的基因,证实了aFGF基因整合到红花基因组中。这为红花作为植物生物反应器的研究奠定了扎实的基础。
OBJECTIVE: To fuse the aFGF with safflower oil body protein and transfer it into safflower. The safflower and aFGF can be double-harvested directly by extracting safflower oil. Methods: Different ages of the leaves of safflower as explants to take orthogonal experimental design method to explore the optimum conditions for safflower regeneration system. Then the aFGF gene was transferred into safflower by agrobacterium-mediated method. The antibiotics of different concentrations were screened and identified by PCR. The positive rate was calculated. Results: According to the different hormonal ratio, the optimal medium for the differentiation of safflower was MS + BA1.0 mg.L-1 + NAA0.2 mg.L-1. The optimal medium for rooting was 1/4 MS + NAA2.0 mg.L-1 + IAA0.5 mg.L-1.3 different types and concentrations of antibacterial antibacterial antibacterial effect of safflower after dipping slightly different, combined with antibacterial effect, pollution rate , Bud differentiation rate and cost issues and other factors, antibacterial antibiotics selection Tim, the concentration of 400 mg.L-1 can inhibit the growth of bacteria, but also conducive to bud differentiation; hyg screening pressure is selected 6 mg.L-1 for its critical concentration; 8 transformed plants were identified, the positive rate was 25%. Conclusion: Through the pathways of organogenesis, the best hormone combination and ratio of safflower differentiation and rooting were determined, and the regeneration system of safflower was established. The optimal concentrations of antibacterial antibiotics and antibiotics were determined. After regeneration, Plants, aFGF gene was amplified by PCR and confirmed that the aFGF gene was integrated into safflower genome. This laid a solid foundation for the research of safflower plant bioreactor.