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首页> 美国卫生研究院文献>3 Biotech >Genotype-independent and enhanced in planta Agrobacterium tumefaciens-mediated genetic transformation of peanut Arachis hypogaea (L.)
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Genotype-independent and enhanced in planta Agrobacterium tumefaciens-mediated genetic transformation of peanut Arachis hypogaea (L.)

机译：基因型非依赖性和植物根癌农杆菌介导的花生遗传转化中的增强花生。

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Agrobacterium infection and regeneration of the putatively transformed plant from the explant remains arduous for some crop species like peanut. Henceforth, a competent and reproducible in planta genetic transformation protocol is established for peanut cv. CO7 by standardizing various factors such as pre-culture duration, acetosyringone concentration, duration of co-cultivation, sonication and vacuum infiltration. In the present investigation, Agrobacterium tumefaciens strain EHA105 harboring the binary vector pCAMBIA1301–bar was used for transformation. The two-stage selection was carried out using 4 and 250 mg l−1 BASTA® to completely eliminate the chimeric and non-transformed plants. The transgene integration into plant genome was evaluated by GUS histochemical assay, polymerase chain reaction (PCR), and Southern blot hybridization. Among the various combinations and concentrations analyzed, highest transformation efficiency was obtained when the 2-day pre-cultured explants were subjected to sonication for 6 min and vacuum infiltrated for 3 min in Agrobacterium suspension, and co-cultivated on MS medium supplemented with 150 µM acetosyringone for 3 days. The fidelity of the standardized in planta transformation method was assessed in five peanut cultivars and all the cultivars responded positively with a transformation efficiency ranging from minimum 31.3% (with cv. CO6) to maximum 38.6% (with cv. TMV7). The in planta transformation method optimized in this study could be beneficial to develop superior peanut cultivars with desirable genetic traits.Electronic supplementary materialThe online version of this article (10.1007/s13205-018-1231-1) contains supplementary material, which is available to authorized users.
机译：农杆菌感染和从外植体推定的转化植物的再生对于诸如花生之类的一些农作物而言仍然艰巨。从此，建立了花生遗传变异的一种能在植物遗传转化中胜任的技术。通过标准化各种因素（例如预培养时间，乙酰丁香酮浓度，共培养时间，超声处理和真空渗透）来实现CO7标准化。在本研究中，使用带有二元载体pCAMBIA1301-bar的根癌农杆菌EHA105进行了转化。使用4和250 mg l -1 BASTA ®进行两阶段选择，以完全消除嵌合和未转化的植物。通过GUS组织化学分析，聚合酶链反应（PCR）和Southern blot杂交评估转基因整合到植物基因组中。在分析的各种组合和浓度中，将2天预培养的外植体在土壤杆菌悬液中超声处理6分钟并真空渗入3分钟，然后在补充有150μM的MS培养基上共培养，可获得最高的转化效率乙酰丁香酮3天。在五个花生品种中评估了植物转化标准方法的保真度，所有品种的转化效率均为正，从最小31.3％（对于Cv。CO6）到最大38.6％（对于Cv.TMV7）。本研究中优化的植物内转化方法可能有益于开发具有理想遗传特性的优良花生品种。电子补充材料本文的在线版本（10.1007 / s13205-018-1231-1）包含补充材料，可以授权使用。用户。

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