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花青素是一种水溶性的黄酮类物质,可使植物呈现出红、蓝、紫和红紫等颜色.外源花青素代谢调控基因的表达可使花青素在植物细胞内积累,使植物体外观上表现出色彩的变化,易于观察,因此可作为报告基因用于植物转基因研究,快速报告细胞、组织、器官或植株是否被转化.本研究用花青素代谢调控基因Bi和C1作为报告基因,以epsp作为筛选标记基因,构建了植物表达载体pBAC9009.基因枪转化玉米自交系501的幼胚,经过除草剂草甘膦抗性筛选和分化再生,获得了转化再生植株75株,其中43株获得结实种子.T0代植株有18株在苗期或生长阶段表现局部或全紫色,8个结实果穗有零星的紫色种子,从外观上可直接确认为转化植株.结合PCR和RT-PCR的分子检测及花青素含量分析,表明叶片和籽粒为紫色的玉米植株中外源目的基因已整合并且高效表达,即紫色表型与分子检测的结果高度一致.该结果表明我们成功建立了以花青素基因作为报告基因的植物转基因可视化跟踪表达系统.该系统的应用可以大大提高工作效率,节约检测成本,对于植物转基因研究具有重要意义.
Anthocyanin is a water-soluble flavonoids, plants can show red, blue, purple and purple and other colors.Exogenous anthocyanin metabolic regulation of gene expression can make anthocyanins accumulate in plant cells, So that the appearance of plants showed a color change, easy to observe, it can be used as a reporter gene in plant gene research to quickly report whether cells, tissues, organs or plants have been transformed.In this study, anthocyanin metabolic regulation genes Bi and C1 As a reporter gene, epsp was used as a screening marker gene to construct a plant expression vector pBAC9009, which was transformed into immature embryos of maize inbred line 501 by in vitro bombardment. After glyphosate resistance screening and differentiation regeneration, 75 transformed plants , Of which 43 obtained strong seeds.On the T0 generation, 18 plants showed partial or full purple in seedling or growth stage, and 8 spikelets had sporadic purple seeds, which could be directly identified as transformed plants by appearance.Combined with PCR and RT -PCR molecular analysis and anthocyanin content analysis showed that the exogenous gene in maize plants with purple leaves and grains was integrated and highly expressed, that is, the height of purple phenotype and molecular detection Cause. The results indicate that we have successfully established a visual tracking system anthocyanin gene expression as a reporter gene in transgenic plants. Application of this system can greatly improve efficiency, saving the cost of testing for gene transfer in plants is important.