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Paper Information

Journal:   IRANIAN JOURNAL OF RANGELANDS AND FORESTS PLANT BREEDING AND GENETIC RESEARCH   spring-summer 2018 , Volume 26 , Number 1 (51) ; Page(s) 1 To 11.

Systematic knockdown of thebaine 6-O-demethylase gene in Papaver somniferum L., using virus-induced gene silencing technique

Author(s):  Alipour Amraei s., ISMAILI A.*, NAZARIAN FIROUZABADI F., LATIFI A.M.
* Department of Agronomy and Plant Breeding, College of Agriculture, Lorestan University, Khorramabad, I.R.Iran
Papaver somniferum is one of the important commercial sources for several pharmaceutical alkaloids including the narcotic analgesics codeine and morphinane and the semi-synthetic drugs oxycodone, buprenorphine and naltrexone. One of the downstream genes in alkaloid biosynthesis pathway in the species is thebaine-6-O demetylase (T6ODM) that inverts thebaine and oripavine to codeinone and morphinone, respectively. In this study, a transient silencing construct based on virus-induced gene silencing (VIGS) technique was used to knockdown the T6ODM expression. In this regard, cDNA of T6ODM was prepared using specific primers and the results of reaction were sequenced for confirmation. Then, a part of resulted cDNA (silencing fragment) was selected for cloning in pTRV vector. The silencing construct was infiltrated to plant leaves via Agrobacterium tumefaciens. The presence of cp (coat proteine) gene of TRV vector was assayed by PCR and the positive TRV plants were screened for analysis of gene expression by semi quantitative and real-time PCR. Results of gene expression analysis showed that transgenic plants had a significant decrease in expression (with average amount of 83. 71%), comparing to control plants.
Keyword(s): Agrobacterium,Papaver,Real time-PCR,VIGS
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