تجزيه و تحليل بيان ژن و توليد آرتميزينين در واكنش به سالسيليك اسيد و جازمونيك اسيد در گياه ترخون﴿Artemisia dracunculus﴾

چكيده انگليسي :

Gene Expression Analysis and Artemisinin Production in Tarragon Artemisia dracunculus in Response to Exogenous Salicylic Acid and Jasmonic Acid Treatment Salimeh Sadeghi Salimeh sadeghi@gmail com 14 June 2015 Department of Agricultural Biotechnology Isfahan University of Technology Isfahan 84156 83111 Iran Degree M Sc Language FarsiSupervisor M Talebi mtalebi@cc iut ac irAbstract Artemisinin produced in very low amounts in Artemisia annua L plants is one of the most effective drugsin treating malaria also has been shown to have selective toxicity towards cancer cells Because its chemicalsynthesis is complicated and economically unfeasible A annua is currently the only source of artemisinin According to previous studies among the Artemisia species the highest concentration of artemisinin reportedfirst in A annua leaves and then in the leaves of A dracunculus Considering that A annua plant is only forwet climates cultivation and proliferation this plant Limitations faced in dry climates but tarragon cultivationis without these limitations Thus increasing the concentration of artemisinin in tarragon plant will result ineasier access and more to this drug material Plant growth regulators PGRs such as Jasmonic acid andSalicylic acid stimulate growth and terpenoid biosynthesis in various aromatic plants and play an importantrole in regulating the reprogramming of gene expression and in eliciting the biosynthesis of secondarymetabolites in plant cells In this study the effect of exogenous applications of Jasmonic acid and Salicylicacid in artemisinin production and the main genes of its biosynthesis pathway such as cpr cyp and dbr2 wereinvestigated For this purpose first to ensure uniformity of studied plants plantlets Proliferated by tissueculture The results of SRAP marker confirmed the genetic stability of plants from tissue culture Plants fromtissue culture treated by 1000 micro molar Salicylic acid and 300 micro molar Jasmonic acid and samplingfrom them were conducted in 0 3 6 12 and 24 hours after treatment Gene expression pattern was monitoredby qRT PCR Technique Then artemisinin content was determined by high performance liquidchromatography HPLC in 0 12 24 48 and 72 hours after treatment The result of the qRT PCRexperiments showed that the Jasmonic acid and Salicylic acid induced significant down regulation of dbr2 inthree hours after treatment but Jasmonic acid and salicylic acid induced significant up regulation of cpr andcyp in six and 24 hours after treatment respectively Previous researches showed that co overexpressing ofcyp and cpr genes could increase the artemisinin content in A annua Overexpressing a single gene is notsufficient to increase artemisinin accumulation because CPR is a redox partner for CYP and helps CYP tocatalyze the conversion of amorphadiene to more oxygenated products in vivo HPLC analysis also showedthat the artemisinin content was increased in 48 hours after Jasmonic acid and 72 hours after Salicylic acidtreatment According to the results of HPLC analysis this study is also proved that co overexpressing of cypand cpr genes could increase the artemisinin content in tarragon Therefore the present study demonstratedthat reduction of dbr2 expression gene had not negative effect on artemisinin biosynthesis since theartemisinic aldehyde is the last common intermediate of the two pathways leading to artemisinin orarteannuin B respectively by DBR2 or CYP enzyme Therefore reducing in content of each them leads toother enzyme Produce artemisinin from one of the two pathways It can be concluded that phyto hormonetreatment can increase some of genes involved in artemisinin biosynthesis pathway and consequently theproduction of artemisinin Keywords Artemisinin Salicylic acid Jasmonic acid HPLC qRT PCR

صادقي، سليمه

تجزيه و تحليل بيان ژن و توليد آرتميزينين در واكنش به سالسيليك اسيد و جازمونيك اسيد در گياه ترخون﴿Artemisia dracunculus﴾

HPLC وqRT - PCR