Optimization of the basal medium for improving production and secretion of taxanes from suspension cell culture of <it>Taxus baccata</it> L

<p>Abstract</p> <p>Background and purpose of the study</p> <p>Taxol is one of the most effective anticancer drugs that isolated from <it>Taxus</it> sp. due to the slow growth of <it>Taxus</it> trees and low concentration of Taxol in the tissues, the biotechnological approaches especially plant cell culture have been considered to produce Taxol in commercial scale.</p> <p>Methods</p> <p>We investigated the effects of basal medium type used in culture media on production of Taxol and other taxane compounds from cell suspension culture of <it>T. baccata</it> L. Briefly, five commonly basal media including Gamborg, Murashige and Skoog, Woody Plant, Schenk and Hildebrandt, and Driver and Kuniyuki medium were used for preparing separate suspension culture media. The intra- and extra-cellular yields of taxanes were analyzed by using HPLC after 21 days period of culturing.</p> <p>Results</p> <p>The yields of taxanes were significantly different for the cultures prepared by different basal media. Moreover, the effects of basal medium on the yield of products differed for varius taxane compounds. Maximum yields of Baccatin III (10.03 mgl^-1) and 10-deacetyl baccatin III (4.2 mgl^-1) were achieved from the DKW basal media, but the yield of Taxol was maximum (16.58 mgl^-1) in the WPM basal media. Furthermore, the secretion of taxanes from the cells into medium was also considerably affected by the type of basal medium. The maximum extra-cellular yield of Taxol (7.81 mgl^-1), Baccatin III (5.0 mgl^-1), and 10-deacetyl baccatin III (1.45 mgl^-1) were also obtained by using DKW basal medium that were significantly higher than those obtained from other culture media.</p

Deep Sequencing Reveals Transcriptome Re-Programming of Cells to the Elicitation with Methyl Jasmonate

Background Plant cell culture represents an alternative source for producing high-value secondary metabolites including paclitaxel (Taxol®), which is mainly produced in Taxus and has been widely used in cancer chemotherapy. The phytohormone methyl jasmonate (MeJA) can significantly increase the production of paclitaxel, which is induced in plants as a secondary metabolite possibly in defense against herbivores and pathogens. In cell culture, MeJA also elicits the accumulation of paclitaxel; however, the mechanism is still largely unknown. Methodology/Principal Findings To obtain insight into the global regulation mechanism of MeJA in the steady state of paclitaxel production (7 days after MeJA addition), especially on paclitaxel biosynthesis, we sequenced the transcriptomes of MeJA-treated and untreated Taxus × media cells and obtained ∼ 32.5 M high quality reads, from which 40,348 unique sequences were obtained by de novo assembly. Expression level analysis indicated that a large number of genes were associated with transcriptional regulation, DNA and histone modification, and MeJA signaling network. All the 29 known genes involved in the biosynthesis of terpenoid backbone and paclitaxel were found with 18 genes showing increased transcript abundance following elicitation of MeJA. The significantly up-regulated changes of 9 genes in paclitaxel biosynthesis were validated by qRT-PCR assays. According to the expression changes and the previously proposed enzyme functions, multiple candidates for the unknown steps in paclitaxel biosynthesis were identified. We also found some genes putatively involved in the transport and degradation of paclitaxel. Potential target prediction of miRNAs indicated that miRNAs may play an important role in the gene expression regulation following the elicitation of MeJA. Conclusions/Significance Our results shed new light on the global regulation mechanism by which MeJA regulates the physiology of Taxus cells and is helpful to understand how MeJA elicits other plant species besides Taxus