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首页> 外文期刊>Molecular biotechnology >A simple method for the production of highly competent cells of Agrobacterium for transformation via electroporation.
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A simple method for the production of highly competent cells of Agrobacterium for transformation via electroporation.

机译：一种生产农杆菌高效能细胞的简单方法，可通过电穿孔进行转化。

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摘要

The introduction of binary plasmids into Agrobacterium hosts for Agrobacterium-mediated transformation of plants is most readily achieved by electroporation. However, occasionally, no transformed colonies are recovered and the transformation program is delayed. Poor transformation rates are commonly associated with particular combinations of Agrobacterium strains and plasmid-selection markers. In order to avoid this problem, it is important for the bacteria to have a highly competent status for reception of plasmid DNA. It is also important to optimize the level of antibiotic for the selection of transformed colonies. In this article, we demonstrate that transformation competence is strongly related to the phase of growth at which a bacterial culture is prepared for electroporation, and we describe a simple procedure that allows the level of transformation-competent cells to be maximized. We have observed that there is significant variation between transformed Agrobacterium strains in the levels of antibiotic tolerance; we define the antibiotic levels that are appropriate for selection of three Argobacterium tumefaciens (EHA101, LBA4404, C58) and two Agrobacterium rhizogenes (LBA9402, Ar2626) strains, transformed with three alternative resistance markers (spectinomycin(res), kanamycin(res), and gentamycin(res)).
机译：通过电穿孔最容易实现将二元质粒引入土壤杆菌宿主以用于土壤杆菌介导的植物转化。但是，有时候，没有回收到转化的菌落，并且转化程序被延迟。转化率低通常与农杆菌菌株和质粒选择标记的特定组合有关。为了避免这个问题，重要的是细菌具有高度能力来接受质粒DNA。优化抗生素水平以选择转化菌落也很重要。在本文中，我们证明了转化能力与细菌培养准备进行电穿孔的生长阶段密切相关，并且我们描述了一种简单的方法，可以使具有转化能力的细胞的水平最大化。我们已经观察到，转化的农杆菌菌株之间在抗生素耐受性水平上存在显着差异。我们定义了适用于选择三种根癌农杆菌（EHA101，LBA4404，C58）和两种发根农杆菌（LBA9402，Ar2626）菌株的抗生素水平，并用三种替代抗性标记（壮观霉素，卡那霉素和庆大霉素）。

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