[Construction of tobacco BY-2 cell lines for heterologous synthesis of taxadiene and transcriptomic analysis].

Paclitaxel is an active anticancer ingredient with extremely high medicinal value, whereas its natural plant resources are scarce. Taxadiene, as the first committed precursor in the biosynthesis of paclitaxel, is of great significance for the production of paclitaxel. This study aimed to establish an efficient heterologous synthesis system for taxadiene, so as to provide a theoretical basis and experimental support for its large-scale production. Using the wild-type tobacco BY-2 cell line and the plant expression vector TS-pk7FGW2, we constructed a tobacco BY-2 cell line heterologously expressing taxadiene synthase (TS) for synthesis of taxadiene. The accumulation of taxadiene in the BY-2 cell line heterologously expressing TS was the highest [0.55 μg/g fresh weight (FW)] under dark culture and reached 1.34 μg/g FW under light treatment, which was 1.4-fold higher than that under dark culture. Transcriptomic analysis was performed on 7-day-old wild-type and TS-expressing tobacco BY-2 cell line to identify differentially expressed genes and associated metabolic pathways. A total of 6 121 differentially expressed genes (DEGs) were identified in the TS-expressing line in comparison with the wild type. Kyoto encyclopedia of genes and genomes (KEGG) enrichment analysis showed that the MVA and MEP pathways in the terpenoid backbone biosynthesis pathway were affected in the tobacco BY-2 cells heterologously expressing TS. This study provides a theoretical basis for optimizing the efficiency of heterologous taxadiene synthesis in BY-2 cells and offers theoretical and data support for elucidating the molecular mechanism by which TS regulates the growth and development of BY-2 cells and the heterologous biosynthesis of taxadiene.