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首页> 外文期刊>Acta Horticulturae >Power and limitations of dicistronic vector systems for plant transformations.
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Power and limitations of dicistronic vector systems for plant transformations.

机译：双顺反子载体系统在植物转化中的功能和局限性。

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摘要

Dicistronic vector constructs harboring an internal ribosomal entry site (IRES) element facilitate the coordinated co-expression of physically independent proteins, as, e.g., a target protein along with a reporter protein. Dicistronic vector constructs have been studied intensively in mammalian cell systems, but their efficiency and applicability in plant cell systems is much less known. The influence of the 1st cistron nucleotide sequence on the translational expression level of the reporter gene encoded by the 2nd cistron was investigated. The testing set consists of the plant viral TMV IRES element and the mammalian viral polio IRES element in combination with either the Atnhx gene, coding for a Na+/H+ antiporter, or the gus gene. Using the resulting four combinations the impact on a firefly luciferase (luc) as reporter gene was studied at the transcriptional, translational and phenotypical level. In order to discriminate between nucleotide sequence of the 1st cistron and its gene product, we introduced a frame shift in the 1st cistron coding sequence. The sequence of the IRES element strongly influenced the translational reporter gene expression present in the 2nd cistron. The results show that the dicistronic vector construct has to be carefully designed and tested for each individual application to obtain the desired level of translational expression of the reporter gene.
机译：带有内部核糖体进入位点（IRES）元件的双顺反子载体构建体促进了物理独立蛋白（例如，靶蛋白和报道蛋白）的协同共表达。双顺反子载体构建体已经在哺乳动物细胞系统中进行了深入研究，但是其在植物细胞系统中的效率和适用性却鲜为人知。研究了第一顺反子核苷酸序列对第二顺反子编码的报道基因翻译表达水平的影响。测试集由植物病毒TMV IRES元件和哺乳动物病毒脊髓灰质炎IRES元件与编码Na + / H + 反转运蛋白的Atnhx基因组合而成，或gus基因。使用所得的四种组合，在转录，翻译和表型水平研究了萤火虫荧光素酶（luc）作为报告基因的影响。为了区分第一顺反子的核苷酸序列及其基因产物，我们在第一顺反子编码序列中引入了移码。 IRES元件的序列强烈影响第二顺反子中存在的翻译报告基因表达。结果表明，必须为每个单独的应用仔细设计和测试双顺反子载体构建体，以获得所需水平的报道基因翻译表达。

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