Taxane diterpenes are secondary metabolites with an important pharmacological role in the treatment of cancer. Taxus spp. biofactories have been used for taxane production, but the lack of knowledge about the taxane biosynthetic pathway and its molecular regulation hinders their optimal function. The difficulties in introducing foreign genes in Taxus spp. genomes hinders the study of the molecular mechanisms involved in taxane production and a new approach is required to overcome them. In this work, a reliable, simple and fast method to obtain Taxus x media protoplasts was developed, allowing their manipulation in downstream assays for the study of physiological changes in Taxus spp. cells. Using this method, Taxus protoplasts were transiently transfected for the first time, corroborating their suitability for transfection assays and the study of specific physiological responses. The two assayed transcription factors (BIS2 and TSAR2) had a positive effect on the expression of several taxane-related genes, suggesting their potential use for the improvement of taxane yields. Furthermore, the results indicate that the developed method is suitable for obtaining Taxus x media protoplasts for transfection with the aim of unravelling regulatory mechanisms in taxane production.

中文翻译：

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转染红豆杉x媒体原生质体以研究转录因子BIS2和TSAR2作为紫杉烷相关基因的激活剂。

紫杉烷二萜是次生代谢产物，在癌症的治疗中具有重要的药理作用。红豆杉属。生物工厂已经用于生产紫杉烷，但是对紫杉烷生物合成途径及其分子调控的了解不足阻碍了它们的最佳功能。在红豆杉属植物中引入外源基因的困难。基因组阻碍了紫杉烷生产中涉及的分子机制的研究，需要一种新的方法来克服它们。在这项工作中，开发了一种可靠，简单，快速的方法来获得Taxus x培养基原生质体，从而可以在下游测定中对其进行操作，以研究Taxus spp的生理变化。细胞。使用这种方法，首次对Taxus原生质体进行了瞬时转染，证实了它们适用于转染测定和特定生理反应的研究。两种测定的转录因子（BIS2和TSAR2）对几种紫杉烷相关基因的表达具有积极影响，表明它们可能用于提高紫杉烷产量。此外，结果表明，所开发的方法适用于获得用于转染的Taxus x培养基原生质体，目的是阐明紫杉烷生产中的调控机制。