Enhanced thebaine and noscapine production and modulated gene expression of tyrosine/dopa decarboxylase and salutaridinol 7-O-acetyltransferase genes in induced autotetraploid seedlings of Papaver bracteatum Lindl

Abstract

Papaver bracteatum, belonging to Papaveraceae, is considered to be a primary ingredient in the production of codeine and anti-drug drugs (e.g., naloxone and Naltrexone) without the involvement of morphine. For polyploidy induction, papaver seedlings were treated with various colchicine concentrations (0, 0.05, 0.1, 0.2 and 0.5% w/v) in three time durations (24, 48 and 72 h). The effects of polyploidy induction on morphological, physiological, alkaloid production and gene expression levels in the pathway of benzylisoquinoline alkaloids were analyzed. High rates of seedling survival were observed at lower colchicine concentrations (0.05 and 0.1% w/v) versus higher concentrations (0.2 and 0.5% w/v). The best polyploidy treatment to induce P. bracteatum shoot tips was observed in plants treated with 0.2% colchicine for 72 h. P. bracteatum-induced tetraploids showed higher stomata and an increase in leaf area (mm2), chlorophyll (a, b and total) and a decrease in stomata density, leaf length (cm) and petiole length. Flow cytometry and chromosome counting analysis revealed that chromosome number was 2n = 2x = 14 in diploid plants and 2n = 4x = 28 in tetraploid plants. Gene expression analysis through semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) showed that the transcription level of sat and tydc genes of the benzylisoquinoline biosynthesis pathway was enhanced in tetraploid plants. The tetraploid plants of P. bracteatum also showed 5.86 and 30.55 times higher thebaine and noscapine contents than the diploid plants, respectively. Regarding the higher commercial value of thebaine and noscapine, tetraploids can efficiently be used to improve the thebaine and noscapine production of P. bracteatum.