In vitro production of secondary metabolite using Atropa komarovii Bline&Shal (Solanaceae) hairy root culture via Agrobacterium rhizogenes ATCC15834

Document Type: Original article

Authors

1 Department of Biology, Science and Research Branch, Islamic Azad University, Tehran, Iran

2 Department of Pharmacognosy, Faculty of Pharmacy, Tehran University of Medicinal Sciences, Tehran, Iran

3 Department of Plant Science, Faculty of Biological Science, Kharazmi University, Tehran, Iran

Abstract

Background & Aim:A new sustainable tissue-based system is presented by plant hairy roots, preserving all of the several specialized types of cell with critical roles in allowing bioactive secondary molecules to be synthesized more consistently as usual. The system is also essential for studying the production of alkaloid in culture.
Experimental: The Atropa komarovii leaves were wounded and infected with soil gram-negative bacterium Agrobacterium rhizogenes ATCC15834. After three weeks, the transformation roots and control roots without infection, appeared, and for confirming that T-DNA Ri plasmid fragments were transformed and integrated to plant genome, the rolB gene region, was amplified using PCR. HPLC method was then used for assaying how two tropane alkaloids such as atropine (hyosciamine) and scopolamine (hyoscine) were produced in hairy roots,control roots, leaves and roots of plantlet.
Results: The data indicated that diagnostic 500bp rol B product amplification was exhibited to be present by all the transformed hairy roots. Scopolamine content in hairy roots was considerably greater than that in control roots but greatest (Hyoscyamine) atropine content was observed in control roots. Analysis of DW, FW and root length showed that fresh and dry root weight increased in hairy roots compared with that in non transformed root.
 Recommended applications/industries: The present study demonstrated that secondary metabolite production using medicinal plants concerns many researchers worldwide today and hairy root culture is a useful method for producing tropane alkaloids in solanaceae.

Keywords


Article Title [Persian]

تولید متابولیت های ثانویه با استفاده از کشت ریشه های موئین در Atropa komarovii از طریق Agrobacerium rhizogenes ATCC15835.

Authors [Persian]

  • افیلا بنی هاشمی 1
  • رمضانعلی خاوری نژاد 1
  • نرگس یاسا 2
  • فرزانه نجفی 3
1 گروه بیولوزی، واحد علوم و تحقیقات، دانشگاه ازاد اسلامی، تهران، ایران
2 گروه فارماکوگنوزی، دانشکده داروسازی، دانشگاه تهران علوم پزشکی تهران، تهران، ایران
3 گروه علوم گیاهی، دانشکده علوم زیستی، دانشگاه خوارزمی، تهران، ایران
Abstract [Persian]

هدف و پیش زمینه: کشت ریشه های موئین روش جدید و سیستمی پایدار بافتی می باشد که برای دخیره نمودن انواع سلول های تخصص یافته که در سنتز و نگهداری مولکول های فعال ثانویه و همچنین نقش مهمی در تحقیق آلکالوئیدها در کشت بافت ایفا می کند.
روش کار:برگ های Atropa komarovii خراش داده شدو توسط باکتری گرم منفی Agrobacterium rhizogenes ATCC15834 آلوده گردید. بعد از سه هفته ریشه های تراریخت و ریشه های شاهد بدون آلودگی ظاهر شدند و برای تصدیق تراریختی و انتفال ناحیه T-DNA از پلاسمید Ri به ژنوم گیاه تکثیر ناحیه ژن rol B توسطPCR صورت گرفت سپس سنتز دو تروپان آلکالوئید اسکوپولامین (هیوسین) و اتروپین (هیوسیامین) در ریشه های موئین و ریشه شاهد و برگ و ریشه حاصل از رشد گیاهچه توسط HPLC ارزیابی گردید.
بحث و نتایج:طلاعات بیان می دارد که تمام ریشه های تراریخت شده حضور ژن rol B 500kbp را نشان دادند. محتوای اسکوپولامین در ریشه های موئین نسبت به ریشه های شاهد به طور قابل ملاحظه ای بالاتر یود و بیشترین میزان آتروپین (هیوسیامین) در ریشه های شاهد دیده شد.
بررسی FW,DW و طول ریشه در دو تیمار ریشه موئین و شاهد نشان داد که میزان وزن تر و وزن خشک همچنین طول ریشه در ریشه های موئین بیشتر یود.
توصیه کاربردی: تحقیق حاضر نشان می دهد که امروزه در دنیا تولید متابولیت های ثانویه از گیاهان دارویی بسیار قابل توجه بوده و کشت ریشه های موئین روش مفید برای تولید تروپان آلکالوئیدها در تیره سیب زمینی می باشد.

Keywords [Persian]

  • Atropa komarovii
  • گیاه دارویی - rolB - تراریختی - تروپان آلکالوئید
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