Comparison of efficiency and time to regeneration of Agrobacterium-mediated transformation methods in Medicago truncatula
Comparison of efficiency and time to regeneration of Agrobacterium-mediated transformation methods in Medicago truncatula
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DOI：
10.1186/s13007-019-0404-1
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发表时间：
2019-02-28
期刊：
影响因子：
5.1
通讯作者：
Frugoli, Julia
中科院分区：
文献类型：
--
作者：
Wen, Li;Chen, Yuanling;Frugoli, Julia
背景植物的组织培养转化具有艺术元素，实验室之间传递协议但通常不直接比较。由于蒺藜苜蓿已作为豆科植物的模型系统而流行，因此快速转化至关重要，并且存在许多方案，但结果各不相同。结果利用蒺藜苜蓿生态型 R108 和 A17 来比较对先前使用的基于芽外植体的方案的修改对愈伤组织产生的转化植物百分比的影响。然后将该百分比与基于 R108 生态型中的根外植体的另外两个转化方案进行比较。分析了胚胎组织来源、培养基成分、转化时间和载体的变化。结论虽然没有获得 A17 转基因植物，但通过在一组条件下对两种广泛研究的生态型进行比较，R108 生态型的转基因植株在短短 4 个月内就产生了。虽然所测试的方案在产生的转化植物的百分比方面给出了相似的结果，但发现所测试的根外植体方案之间对转基因的劳动力和时间的考虑有所不同。这些考虑因素可能会影响选择哪种方案来引入单个转基因，还是使用 CRISPR/Cas9 构建体创建具有多个突变的品系。
BackgroundTissue culture transformation of plants has an element of art to it, with protocols passed on between labs but often not directly compared. As Medicago truncatula has become popular as a model system for legumes, rapid transformation is critical, and many protocols exist, with varying results.ResultsThe M. truncatula ecotypes, R108 and A17, were utilized to compare the effect of a modification to a previously used protocol based on shoot explants on the percentage of transformed plants produced from calli. This percentage was then compared to that of two additional transformation protocols based on root explants in the R108 ecotype. Variations in embryonic tissue sources, media components, time for transformation, and vectors were analyzed.ConclusionsWhile no A17 transgenic plants were obtained, transgenic plantlets from the R108 ecotype were produced in as little as 4 months with a comparison of the two widely studied ecotypes under a single set of conditions. While the protocols tested gave similar results in percentage of transformed plants produced, considerations of labor and time to transgenics that vary between the root explant protocols tested were discovered. These considerations may influence which protocol to choose for introducing a single transgene versus creating lines with multiple mutations utilizing a CRISPR/Cas9 construct.