تحريك ريشه‌هاي موئين تراريخت در گياه دارويي شقايق (Papaver somniferum L) با استفاده از آگروباكتريوم رايزوژنز (Agrobacterium rhizogenes)

گياه شقايق (Papaver somniferum L.) يكي از قديمي‌ترين گياهان داروئي شناخته شده است. با توجه به اين‌كه پتانسيل مواد دارويي گياهان دارويي درشرايط طبيعي بسيار محدود مي‌باشد، از اين رو كشت بافت گياهي به‌عنوان يكي از روش‌هاي كاربردي نقش مهمي در راستاي توليدصنعتي متابوليت‌هاي ثانويه درگياهان دارويي دارد. از ميان سيستم‌هاي كشت بافت، سيستم ريشه‌هاي موئين درشرايط كشت بدون تنظيم‌كننده رشد بسيار پايدار است و به دليل ثبات ژنتيكي و بيوشيميايي ذاتي، متابوليت‌هاي ثانويه را در طي يك دوره‌ي طولاني توليد مي‌كنند. در اين پژوهش به منظور بهينه‌سازي شرايط كشت ريشه‌هاي موئين از سويه‌هاي مختلف آگروباكتريوم رايزوژنز (ATCC15834،A4 و GMI9534)، محيط‌ هاي كشت MS و ½MS، دو دماي 25 و 17 درجه سلسيوس در دوره هم‌كشتي و دو نوع ريزنمونه‌ي اندام هوايي و محور زيرلپه استفاده گرديد. صفت‌هاي درصد تحريك ريشه موئين، تعداد ريشه موئين در ريزنمونه و تعداد انشعاب فرعي ريشه موئين در طول يك سانتي‌متر از ريزنمونه اندازه‌گيري شد. واكاوي نتايج نشان داد كه سويه‌ي ATCC15834، محيط كشت ½ MS، دماي 25 درجه سلسيوس همكشتي و ريزنمونه اندام هوايي بهترين تركيب تيماري جهت تحريك ريشه موئين در گياه شقايق هستند. با استفاده از واكنش PCR تأييد تراريختي ريشه‌هاي موئين با آغازگرهاي اختصاصي ژن rolB انجام گرفت.

Background and Objectives Opium poppy plant (Papaver somniferum L.) is one of the oldest known medicinal plants. Since the potential of medicinal components production in medicinal plants is very limited in natural conditions, plant tissue culture, as a new technology, has an important role in industrial production of secondary metabolites in medicinal plants. Among different tissue culture systems, hairy root is very stable in hormone-free culture conditions and these roots stably produce secondary metabolites over a long period due to their inherent genetic and biochemical stability. Hence, the objective of this study was to evaluate the effect of some different parameters on optimization of hairy production in P. somniferum. Materials and methods In the present research, for the optimization of hairy root in papaver, a factorial experiment was conducted based on a completely randomized design with 4 replications. The factors included different Agrobacterium rhizogenes strains (ATCC15834, A4 and GMI9534), medium (MS and ½ MS medium), two temperatures (17 and 25 °C) at co-cultivation period, and two different explants (excised shoot and hypocotyl). Different traits were evaluated including percentage of induced hairy roots, number of lateral branches/1cm, number of hairy root per explants and days after inoculation (DAI). Results Hairy roots were induced at the wound site of explants after weeks of culture. Analysis of the results showed that strain ATCC15834, ½ MS medium, temperature 25 °C and excised shoot explant were the best treatment combination for hairy roots induction in the poppy plant. Transformed hairy roots were confirmed by polymerase chain reaction (PCR) using rolB gene specific primers. Discussion All of the A. rhizogenes strains led to hairy root induction, but there were no hairy roots formed from control explants. Differences in virulence, morphology and growth rate can be partially explained by the variety of plasmids harbored by the A. rhizogenesis strains. In the present study, excised shoot explants were highly and significantly susceptible to infection by each strain of A. rhizogenes and hypocotyl explants showed low rate of hairy root induction.