ZHANG Fu-li,SHU Wen-tao,ZHANG Yi,et al.Cluster Bud Induction of Soybean Whole Cotyledonary Node and Application in Agrobacterium mediated Genetic Transformation[J].Soybean Science,2012,31(06):865-868.[doi:10.3969/j.issn.1000-9841.2012.06.001]
大豆整体子叶节再生体系的建立及应用于农杆菌介导遗传转化初探
- Title:
- Cluster Bud Induction of Soybean Whole Cotyledonary Node and Application in Agrobacterium mediated Genetic Transformation
- 文章编号:
- 1000-9841（2012）06-0865-04
- Keywords:
- Soybean; Whole cotyledonary node; Regeneration; Agrobacterium mediated; Genetic transformation
- 分类号:
- S565.1
- 文献标志码:
- A
- 摘要:
- 以中黄20、周豆02005、周豆11和周豆18整体子叶节为材料，以MSB为基本培养基，通过L9(34)正交设计研究了6-BA、IBA和KT 9种浓度组合对大豆整体子叶节丛生芽再生的影响；并将大豆的整体子叶节应用于农杆菌介导的遗传转化研究。结果表明：9种激素组合中以MSB+3.0 mg·L-16-BA+0.1 mg.L-1IBA +0.5 mg.L-1KT最适合周豆02005丛生芽的诱导，而MSB+3.5 mg.L-16-BA+0.2 mg.L-1IBA +0.2 mg.L-1KT最适宜周豆11、周豆18和中黄20的丛生芽诱导；4个大豆基因型中，以中黄20的丛生芽数最多；丛生芽用于诱导生根时，IBA为0.8 mg.L-1、NAA为0.8~1.0 mg.L-1时较适合大豆生根，其中以中黄20生根数最多。以中黄20整体子叶节为材料，进行了农杆菌介导的遗传转化研究。PCR分析和GUS染色初步表明，外源基因转入了大豆的基因组中，转化率为7.8%，得到转基因植株的周期为35~42 d。
- Abstract:
- The effects of nine concentration combinations of 6BA,IBA and KT designed by orthogonal design[L9(34)]were evaluated on induction of soybean[Glycine max(L.)Merrill\]multi shoot from whole cotyledonary nodes.Four soybean genotypes,including Zhouhuang 20,Zhoudou 02005,Zhoudou 11 and Zhoudou 18,were used as plant materials,while MSB was used as basic medium.The results showed that nine concentration combinations had different effects on induction of multi shoot for soybean whole cotyledonary nodes.The regeneration system was further used to Agrobacterium mediated genetic transformation.For nine concentration combinations,MSB+3.0 mg.L-16-BA+0.1 mg.L-1IBA+0.5 mg.L-1KT for Zhoudou 02005 and MSB+3.5 mg.L-16-BA+0.2 mg.L-1IBA+0.2 mg.L-1KT for Zhouhuang 20,Zhoudou 11 and Zhoudou18 were optimum to induce multi shoot than other combinations.For four genotypes,the regeneration efficiency of multi shoot for Zhouhuang 20 was the highest.The concentration of 0.8 mg.L-1 for IBA and 0.8-1.0 mg.L-1for NAA was much more suitable for soybean rooting,and rooting efficiency of Zhonghuang 20 was the highest. Agrobacterium mediated transformation of Zhonghuang 20 was performed using whole cotyledonary nodes as explants,it showed that reporter gene(GUS)had been transformed into soybean genome by PCR analysis and GUS staining with 7.8% transformation efficiency,and the period of obtaining transgenic plant was 35-42 d.
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备注/Memo
基金项目：转基因生物新品种培育重大专项子任务(2008ZX08010-004)；河南省教育厅自然科学项目（2012B210028，2011B180060）；周口师范学院博士科研启动基金；周口师范学院实验室开放项目基金。