نوع مقاله : مقاله کامل پژوهشی

نویسنده

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چکیده

ویروس های زردی غلات از مخرب ترین ویروس های غلات در سراسر دنیا هستند. در این تحقیق پروتئین های P0، سرکوبگرهای خاموشی، در دو ویروس کوتولگی زرد غلات، Cereal yellow dwarf virus-RPV (CYDV-RPV) وCereal yellow dwarf virus- RPS (CYDV-RPS) از نظر محل تجمع و قرارگیری در سلول های گیاه توتون مورد بررسی قرار گرفتند. بدین منظور ترادف ژن های P0 پس از تکثیر در آزمون زنجیره ای پلیمراز (PCR)، با روش Gateway همسانه سازی شدند. سپس سازه های این ژن ها در اتصال با ژن کد کننده green fluorescent protein (GFP) وارد باکتری Agrobacterium tumefaciens شدند و با تزریق باکتری حامل این سازه ها به گیاه به صورت موقتی بیان شدند. پس از 24 ساعت از زمان تزریق، محل قرارگیری این پروتئین ها در سلول های گیاه توسط میکروسکوپ کانفوکال (confocal) مورد بررسی قرار گرفت. نتایج نشان داد که محل تجمع پروتئین P0 در هر دو ویروس در هسته و سیتوپلاسم است اما تمایل آنها برای تجمع متفاوت است. به عبارت دیگر پروتئین P0 در ویروس RPV CYDV- در مقایسه با CYDV-RPS تمایل کمتری برای تجمع در هسته دارد در حالی که پروتئین P0 در ویروس CYDV-RPS بیشتر در هسته متمرکز است. در آزمون لکه برداری وسترن با استفاده از آنتی بادی پلی کلونال GFP، پروتئین های GFP:P0 ردیابی شدند ولی پروتئین GFP به تنهایی دیده نشد که نشان می دهد محل های تجمع دیده شده مربوط به پروتئین های GFP:P0 است.

کلیدواژه‌ها

عنوان مقاله [English]

Subcellular localization of silencing suppressor proteins encoded by two cereal yellow dwarf viruses

نویسنده [English]

  • R. Almasi

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چکیده [English]

Cereal yellow dwarf viruses are among the most destructive cereal viruses worldwide. In this study, subcellular localizations of silencing suppressor proteins ( P0) from two cereal yellow dwarf viruses, Cereal yellow dwarf virus-RPV (CYDV-RPV) and Cereal yellow dwarf virus-RPS (CYDV-RPS), were examined via confocal microscopy. To do so, P0 genes were synthesized by polymerase chain reaction (PCR) and were cloned through Gateway cloning system. Agrobacterium. tumefaciens clones harboring GFP:P0 constructs were infiltrated into Nicotiana benthamiana plants. Transient expression of P0 proteins with N-terminal GPF fusion enabled us to examine their subcellular localizations in plant cells, 24 hours post infiltration. Results showed that both proteins are nuclear-cytoplasmic but different tendencies were observed in their localizations. In other words, P0 of CYDV-RPS was more nuclear oriented than P0 of CYDV-RPV. In the Western blot analysis using GFP polyclonal antibody only GFP:P0 proteins but no free GFP, were detected indicating that localizations were related to GFP:P0 proteins.

کلیدواژه‌ها [English]

  • Polerovirus
  • P0 protein
  • Gateway cloning
  • Western blot
  • subcellular localization
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