The parasitic plant genus Cuscuta is notoriously difficult to transform and to propagate or regenerate in vitro. With it being a substantial threat to many agroecosystems, techniques allowing functional analysis of gene products involved in host interaction and infection mechanisms are, however, in high demand. We set out to explore whether Agrobacterium‐mediated transformation of different plant parts can provide efficient alternatives to the currently scarce and inefficient protocols for transgene expression in Cuscuta. We used fluorescent protein genes on the T‐DNA as markers for transformation efficiency and transformation stability. As a result, we present a novel highly efficient transformation protocol for Cuscuta reflexa cells that exploits the propensity of the infection organ to take up and express transgenes with the T‐DNA. Both, Agrobacterium rhizogenes and Agrobacterium tumefaciens carrying binary transformation vectors with reporter fluorochromes yielded high numbers of transformation events. An overwhelming majority of transformed cells were observed in the cell layer below the adhesive disk’s epidermis, suggesting that these cells are particularly susceptible to infection. Cotransformation of these cells happens frequently when Agrobacterium strains carrying different constructs are applied together. Explants containing transformed tissue expressed the fluorescent markers in in vitro culture for several weeks, offering a future possibility for development of transformed cells into callus. These results are discussed with respect to the future potential of this technique and with respect to the special characteristics of the infection organ that may explain its competence to take up the foreign DNA.

中文翻译：

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基于人工诱导的感染部位的高效转化Cuscuta reflexa的协议。

寄生植物属菟丝子是极难变换和传播或体外再生。由于它对许多农业生态系统构成重大威胁，因此迫切需要能够对参与宿主相互作用和感染机制的基因产物进行功能分析的技术。我们着手探讨农杆菌介导的不同植物部分的转化是否可以提供有效的替代方法，以替代目前稀缺且效率低下的葫芦科转基因表达方案。我们使用T‐DNA上的荧光蛋白基因作为转化效率和转化稳定性的标记。结果，我们提出了一种新的高效反射法，用于Cuscuta reflexa利用感染器官倾向于利用T‐DNA吸收并表达转基因的细胞。这两个，发根农杆菌和农杆菌携带二进制转化载体与记者荧光产生高的数字转化事件。在粘合盘表皮下的细胞层中观察到绝大多数转化细胞，表明这些细胞特别容易感染。当农杆菌属时，这些细胞的共转化频繁发生携带不同构建体的菌株一起应用。含有转化组织的外植体在体外培养数周后表达了荧光标记，为将转化细胞发展成愈伤组织提供了未来的可能性。就该技术的未来潜力以及感染器官的特殊特征（可能解释其吸收外源DNA的能力）进行了讨论。