بررسی چند شکلی DNA در جمعیت‌های AG-4 Rhizoctonia solani با استفاده از نشانگر مولکولی rDNA RFLP

دو فصلنامه تحقیقات بیماریهای گیاهی

مقاله 2، دوره 5، شماره 1، خرداد و تیر 1396، صفحه 11-30   

نوع مقاله: مقاله پژوهشی

نویسندگان

فرزانه بادپا^{* 1}؛ غلامرضا بلالی²؛ بهرام شریف نبی³

¹دانش آموخته کارشناسی ارشد، گروه زیست شناسی، دانشگاه اصفهان، اصفهان، ایران.

²دانشیار، گروه زیست شناسی، دانشکده علوم، دانشگاه اصفهان، اصفهان، ایران.

³استاد، گروه گیاهپزشکی، دانشکده کشاورزی، دانشگاه صنعتی اصفهان، اصفهان، ایران

تاریخ دریافت: 11 بهمن 1395،  تاریخ پذیرش: 11 بهمن 1395 

چکیده

توالی­های DNA ریبوزومی به­دلیل اینکه دارای کپی­های زیادی از ژن­های rRNA و نیز درجه بالایی از تغییر می­باشند، به­منظور بررسی روابط فیلوژنی در محدوده وسیعی از سطوح تاکسونومی مورد استفاده قرار می­گیرند. نواحی ITS و فواصل داخل ژنی DNA ریبوزومی هسته­ای، واحد­های تکراری تکامل یافته ثابت می­باشند و ممکن است در بین گونه های داخل یک جنس یا در بین افراد یک جمعیت متغیر باشند. به­منظور ارزیابی چند شکلی نواحی حد فاصل ژن­های s28 و s18 در ژنوم قارچ Rhizoctonia solani  گروه آناستوموزی 4 (AG-4)، 28جدایه از میزبان­های مختلف مورد ارزیابی قرار گرفتند. از جدایه­ها DNA استخراج و از آن­ها به عنوان الگو در واکنش­های PCR استفاده شد. در تکثیر نواحی فاصله­ساز داخلی بین ژن­های ریبوزومی با استفاده از آغازگر­های ITS1 و ITS4 یک قطعه DNA به اندازه 700 جفت باز ردیابی شد. قطعه مذکور با آنزیم­های برشیTaq I  Xba I, Hae III, Bam HI, Hind III, Sac I, Pst I, Nde I, Xho I, Hinc II, Hinf I, Eco RI, هضم شد. ولی آنزیم­های Xba I, Bam HI, Hind III, Sac I, Pst I, Nde I, Xho I فاقد محل برش بودند. نتایج حاصله نشان داد که جدایه­های Rhizoctonia solani AG4 هتروژن بوده و ITS-RFLP با استفاده از آنزیم Hinc II تفاوت بین زیرگروه­های AG4-HG I و AG4-HG II رااز نظر الگوی برش آنزیمی نشان داد.

کلیدواژه‌ها

تنوع ژنتیکی؛ نواحی فاصله‌ ساز داخلی(ITS)؛ واکنش زنجیره‌ای پلی‌مراز؛ Rhizoctonia solani گروه آناستوموزی 4

عنوان مقاله [English]

Study on DNA polymorphism in populations of Rhizoctonia solani anastomosis group 4 using rDNA RFLP

نویسندگان [English]

F. Badpa¹؛ G.R. Balali²؛ B. Sharifnabi³

چکیده [English]

Ribosomal DNA (rDNA) sequences have been widely used to study the phylogenetic relationships in different fungi. Fungal nuclear rRNA genes are arranged as tandem repeats with several hundred copies per genome. These spacer regions are considerably more variable than the subunit sequences and have been widely used in studies on the relationships among species within a single genus or among intraspecific populations. To evaluate the polymorphism between 18S and 28S genes, 28 isolates of Rhizoctonia solani anastomosis group 4 were examined. Genomic DNA was extracted from the isolates and prepared for PCR reaction. The amplification was performed using internal transcribed spacer (ITS) ITS1 and ITS4 primers. A DNA fragment of 700 bp in size was detected in PCR products of all tested isolates. To assess existence of any further polymorphism in the ITS region, the PCR products were digested with restriction endonucleases. Although there were restriction sites for XbaI, HaeIII, BamHI, HindIII, SacI, PstI, and TaqI endonucleases, there were no restriction sites for NdeI, XhoI, HincII, HinfI, EcoRI and XhoI endonucleases. The endonuclease HincII recognized a restriction site on PCR products that discriminated the isolates belonging to AG4-HGII form isolates of AG4-HGI. Based on the results, it has been concluded that AG-4 isolates of Rhizoctonia solani wereheterogenic.
 

کلیدواژه‌ها [English]

genetic diversity, ITS-rDNA, PCR, Rhizoctonia solani AG4

مراجع

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