Enhanced Production of Thebaine in Cell Suspension Cultures of Papaver bracteatum by overexpression of SalAT gene

The Iranian poppy by the scientific name of Papaver bracteatum Lindl. has been long propounded as a source for thebaine. Thebaine is among secondary metabolites that due to its wide spread and unique medicinal properties play a valuable role in production of herbal medicines and pharmaceutical industries on the whole.The objective of the present study is to investigate the enhanced production of thebaine by over expression of salutaridinol ۷-o-acetyltransferase (SalAT) gene, a key gene in thebaine biosynthesis pathway, in Agorobacterium-mediated transformed cell suspension culture of P. bracteatum. In this study, the cDNA of SalAT with P. bracteatum origin was cloned in pBI۱۲۱ expression vector under CaMV۳۵s promoter. In order to transfer the cloned gene Agrobacterium tumefaciens of LBA۴۴۰۴ strain and hypocotyl–cotyledon explants were used. For recognition of transformed calli obtained, PCR and Southern blot analysis were employed and high performance liquid chromatography analysis (HPLC) was applied for determination of thebaine content. The comparison of callus induction and thebaine content between transformed and non-transformed cell culture was studied on completely randomized design with three replications. According to HPLC analysis which revealed a significant ۱.۷ fold (۰.۵۸ mg/g) enhancement in thebaine content in transformed suspension cell culture comparing to non-transformed cell culture (۰.۳۴ mg/g), it can be concluded that the over expression of key genes such as SalAT gene in the cellular suspension culture of P. bracteatum could be a promising potential for mass production of this valuable alkaloid.

Keywords:

Cell suspension culture , Over expression , Papaver bracteatum , SalAT gene , thebaine