Laboratorio de Fisiología Vegetal, Facultad de Farmacia, Universidad de Barcelona, Joan XXIII s/n, Barcelona, Spain.
Biotechnol Prog. 2010 Jul-Aug;26(4):1145-53. doi: 10.1002/btpr.424.
Dedifferentiated Taxus media cell cultures presenting the same genetic characteristics as the parent culture were established from transformed roots. Two transformed cell lines were studied: Rol C, carrying the T-DNA of A. rhizogenes 9,402 and TXS, carrying both the T-DNA of A. rhizogenes and the txs transgene of T. baccata under the control of the 35S CaMV promoter. In the second part of a previously optimized two-stage system, the transformed cell lines were cultured in a production medium supplemented with the elicitor methyl jasmonate. Taxane production in the transformed cultures was compared with an untransformed T. media cell line cultured in the same conditions. The highest taxane production was observed in the TXS cell line when cultured in the optimized production medium with methyl jasmonate, being 265% greater than in the untransformed control and 170% greater than in the Rol C cell line. However, txs expression and the activity of the enzyme taxadiene synthase in the TXS cells were lower than in the line carrying only the rol genes (Rol C). It is also noteworthy that the taxane production as well as the txs gene expression and TXS activity in all the cell lines, both transformed and untransformed, were clearly dependent on the elicitor action.
从转化的根部分离出具有与亲本培养物相同遗传特征的去分化短叶紫杉细胞培养物。研究了两个转化细胞系：携带发根农杆菌 9,402 T-DNA 的 Rol C 和携带发根农杆菌 T-DNA 和欧洲云杉 txs 转基因的 TXS，后者受 35S CaMV 启动子控制。在之前优化的两阶段系统的第二部分中，将转化细胞系在补充有诱导剂茉莉酸甲酯的生产培养基中进行培养。将转化培养物与在相同条件下培养的未转化短叶紫杉细胞系进行了紫杉醇生产的比较。当在优化的含茉莉酸甲酯的生产培养基中培养时，TXS 细胞系的紫杉醇产量最高，比未转化的对照高 265%，比 Rol C 细胞系高 170%。然而，TXS 细胞中的 txs 表达和 taxadiene 合酶的活性低于仅携带 rol 基因的细胞系（Rol C）。值得注意的是，所有转化和未转化的细胞系中的紫杉醇产量以及 txs 基因表达和 TXS 活性均明显依赖于诱导剂的作用。
