Production and gene expression of morphinan alkaloids in hairy root culture of Papaver orientale L. using abiotic elicitors

Abstract

Nowadays, hairy root culture elicitation is an excellent platform with many advantages which make it an attractive system for using in plant biochemistry and molecular biology. Here, an efficient system for A. rhizogenes transformation in oriental poppy, Papaver orientale, was developed. Meanwhile, the expression level of various genes (COR, SalAT, SalR, T6ODM, CODM and Salsyn) in the morphinan pathway for response to methyl jasmonate (MJ) and salicylic acid (SA) at different time points (6, 12, 24 and 48 h) was examined. Afterwards, in vitro production of thebaine, morphine and codeine was investigated. The wild-type A. rhizogenes strain ATCC15834 was able to produce transformed roots with the highest frequency on the hypocotyl explant. Quantitative reverse-transcription PCR analysis showed that the expression of SalR, Salsyn, SalAT, T6ODM and CODM were significantly up-regulated with MJ treatment at 48 h. Moreover, the expression levels of Salsyn, T6ODM and CODM was up-regulated considerably in SA at 48 h treatment, while, the expression of SalR was down-regulated at 24 h. Furthermore, high-performance liquid chromatography analysis showed that MJ elicitation enhanced thebaine and morphine at 48 h by 2.63-fold (3.08 mg g−1) and 6.18-fold (5.38 mg g−1), respectively, and also codeine was increased at 24 h by 3.67-fold (2.57 mg g−1). In addition, the accumulation of morphine and thebaine was increased by 4.22-fold (2.87 mg g−1) and 2-fold (1.66 mg g−1) in comparison with control treatment after SA treatment for 48 h. Our findings suggest that oriental poppy hairy roots can be used as a good source for producing commercial alkaloids.