Systematic silencing of codeine O-demethylase (CODM) gene, using virus-induced gene silencing (VIGS) technique in Papaver somniferum L

Papaver somniferum L. is the most important commercial source of production of morphinan alkaloid component, such as codeine, thebaine, narcotine and papaverine that are exploited by the pharmaceutical industry as analgesics, antitussives and anti-spasmodics. Recently, metabolic engineering technology for genetic manipulation of alkaloid biosynthesis and metabolic products has been used via production of transgenic plants. Codeine O-demethylase (CODM) catalyzes the late step of morphine biosynthesis in opium poppy for conversion of thebaine to oripavine and codeinone and codeine to morphine via demethylation mechanism. This study was conducted in silencing of CODM gene using VIGS technique. cDNA was amplified using specific primers, and then a part of this cDNA at the 3́ UTR terminal was subcloned in TRV2 vector in Agrobacterium as silencing fragment. Plant leaves was inoculated via Agrobacterium containing silencing fragment. Initial screening of transformed plants was conducted via PCR on tobacco rattle viral coat protein gene (TRV-CP) and then, some positive TRV plants were analyzed using semi-quantitative RT PCR. The final analysis of VIGS was performed using real-time PCR. Based on the results 95 percent reduction in gene expression of CODM in transgenic plants compared to the control plants was achieved.