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بهینه‌سازی روش استخراجRNA از بافت پوست شاخه درختان خانواده مخروطیان | ||
| تحقیقات ژنتیک و اصلاح گیاهان مرتعی و جنگلی ایران | ||
| مقاله 9، دوره 24، شماره 1، اردیبهشت 1395، صفحه 102-113 اصل مقاله (253.09 K) | ||
| نوع مقاله: مقاله علمی - پژوهشی | ||
| شناسه دیجیتال (DOI): 10.22092/ijrfpbgr.2016.106171 | ||
| نویسندگان | ||
| فائزه حائری زاده1؛ اسد معصومی اصل* 2؛ پژمان آزادی3؛ حمید رجبی معماری4؛ نفیسه ابوفاضلی5 | ||
| 1دانشجوی کارشناسی ارشد، اصلاح نباتات، دانشگاه یاسوج، یاسوج | ||
| 2نویسنده مسئول مکاتبات، استادیار، دانشکده کشاورزی، دانشگاه یاسوج، یاسوج پست الکترونیک: Masoumiasl@yu.ac.ir | ||
| 3استادیار، پژوهشکده بیوتکنولوژی کشاورزی ایران، کرج | ||
| 4استادیار، دانشکده کشاورزی، دانشگاه شهید چمران، اهواز | ||
| 5عضو شرکت دانش بنیان زیست فناوران نوین گیتی ژن، تهران | ||
| چکیده | ||
| فرآیند استخراج RNA از بافتهای سخت چوبی و بهطور خاص درختان خانواده مخروطیان بهدلیل داشتن غلظتهای بالایی از ترکیبات فنولیک، پلیساکاریدها، متابولیتهای ثانویه و لیگنین، بسیار مشکل است و تا بهحال هیچگونه کیت تجاری برای این منظور ساخته نشده است. در این مطالعه مشکلات فوق برطرف گردیده و روشی سریع، ساده و اقتصادی برای استخراج RNA از بافت ساقه گونه‌های Cedrus deodara و Abies grandis که هر دو از خانواده مخروطیان میباشند، ارائه شده است. نسبت جذب نوری در طول موج‌های 260 به 280 و 260 به 230 نانومتر، بهترتیب 88/1 و 1/1 برای گونه Cedrus deodara و 8/1 و 9/0 برای گونه Abies grandis بهدست آمد. تغییرات صورت گرفته در جهت بهینه‌سازی استخراج RNA نیز شامل استفاده از تیمار پیش‌گرما روی بافر استخراج، کاهش دور سانتریفیوژ حذف آلودگی DNA و حذف DEPC می‌باشد. ساخت cDNA با استفاده از RNA استخراج شده با موفقیت انجام و واکنش RT-PCR باندهای مورد انتظار را تولید نمود. لذا بهنظر میرسد این روش برای استخراج RNA از بافتهای چوبی مناسب است. | ||
| کلیدواژه‌ها | ||
| بافت چوبی؛ ترکیبات فنولیک؛ درخت نراد؛ DEPC | ||
| عنوان مقاله [English] | ||
| Optimization of RNA extraction protocol for trees bark tissue of coniferous family | ||
| نویسندگان [English] | ||
| faeezeh haeerizadeh1؛ asad masoomi asl2؛ pezhman azadi3؛ hamid rajabi-memari4؛ nafiseh aboofazeli5 | ||
| چکیده [English] | ||
| RNA extraction from hard wood tissues, especially in coniferous family trees, is very hard because they have high concentrations of phenolic compounds, polysaccharides, secondary metabolites and lignin. Moreover, up to now, no commercial kit is made for the mentioned purpose. In this study, the problems were resolved and a rapid, simple, and economic protocol is presented for RNA extraction from bark tissue of Cedrus deodara and Abies grandis, which belong to conifer family. Ratios of optical density of 260/280 nm and 260/230 nm were respectively obtained as 1.88 and 1.1 for Cedrus deodara and 1.8 and 0.9 for Abies grandis. Modifications for optimization of RNA extraction were preheating treatment of extraction buffer, and decreasing centrifuge rounds to eliminate DNA contaminations and DEPC. Synthesis of cDNA was performed using extracted RNA. RT-PCR successfully produced expected bands. Therefore, it seems that the presented protocol is suitable for RNA extraction from woody tissues. | ||
| کلیدواژه‌ها [English] | ||
| Abies grandis, bark tissue, DEPC, Phenolic compounds | ||
| مراجع | ||
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