بارکد گذاری DNA برخی از گیاهان دارویی

نوع مقاله: علمی پژوهشی

نویسندگان

1 کارشناسی ارشد بیوتکنولوژی در کشاورزی، دانشکده علوم کشاورزی، دانشگاه محقق اردبیلی، اردبیل

2 استادیار گروه زراعت و اصلاح نباتات، دانشکده علوم کشاورزی، دانشگاه محقق اردبیلی

3 محقق بیوتکنولوژی گیاهی، گروه اصلاح نباتات، دانشگاه واخنینگن، هلند

4 دانشیار گروه گیاه پزشکی، دانشکده علوم کشاورزی، دانشگاه محقق اردبیلی

چکیده

بارکدگذاری DNA روشی ساده برای شناسایی گونه‌ها با استفاده از یک توالی کوتاه ژنتیکی از یک بخش استاندارد ژنوم است. این روش به منظور شناسایی هشت گیاه دارویی جمع‌آوری‌شده از استان اردبیل مورد استفاده واقع شد. استخراج DNA به روش تغییر‌یافته CTAB انجام گرفت و PCR با آغازگرهای طراحی‌شده بر‌ اساس بارکدهای کلروپلاستی rbcL، trnH-psbA، matK و بارکد هسته‌ای ITS انجام شد. سپس محصولات PCR خالص‌سازی و تعیین توالی گردید. درصد موفقیت تکثیر و توالی‌یابی در نمونه‌ها به‌ترتیب 87 و 62، 75 و 37، 62 و 12، 75 و 37 به‌دست‌آمد. توالی‌ها با نمونه‌های موجود در پایگاه داده NCBI همردیف‌شده و تحت تجزیه بیوانفورماتیکی قرار گرفتند. در درخت خویشاوندی گونه‌های هم‌جنس بر ‌اساس توالی‌های بارکد‌های rbcL و trnH-psbA از دیگر جنس‌ها تفکیک شدند. همچنین، در بارکد ITS فقط شیرین‌بیان با گیاهان هم‌جنس خود قرار گرفت. در این مطالعه، گیاه سوسن‌چلچراغ برای اولین بار با بارکد rbcL بارکدگذاری شد. SNP بارکدهای rbcL (کم‌تر از 30)، trnH-psbA (کم‌تر از 100)، ITS (بیشتر از 200) و matK (کمتر از 20) شمارش شد. بنابراین، بارکد rbcL به‌دلیل قدرت تفکیک بالا، تعداد SNP پایین و جامعیت در اکثر گونه‌ها، به‌عنوان بهترین بارکد معرفی ‌شد. باوجوداین، بارکدهای ITS و trnH-psbA به‌دلیل مشکل مرتبط با توالی‌یابی مستقیم محصولات PCR و عدم دسترسی به توالی‌های با کیفیت، به‌عنوان بارکدهای مکمل شناسایی شدند. بارکد matK به‌دلیل قدرت تکثیر و توالی‌یابی پایین برای نمونه‌های مورد بررسی توصیه نمی‌شود.

کلیدواژه‌ها

موضوعات


عنوان مقاله [English]

DNA barcoding of some local medicinal plants of Ardabil province

نویسندگان [English]

  • Fatemeh Asadi 1
  • Sara Dezhsetan 2
  • Robab Ghahramanzadeh 3
  • Jabraeil Razmjou 4
  • Mohammad Taghi Alebrahim 2
1 M.Sc. Agricultural Biotechnology, University of Mohaghegh Ardabili, Ardabil, Iran.
2 Assistant Professor, Department of Agronomy & Plant Breeding, Faculty of Agricultural Sciences, University of Mohaghegh Ardabili, Ardabil, Iran
3 Plant biotechnology researcher, Plant breeding department, Wageningen UR.
4 Associate Professor, Department of Plant Protection, Faculty of Agricultural Sciences, University of Mohaghegh Ardabili, Ardabil, Iran
چکیده [English]

DNA barcoding is a simple way to identify species using a very short genetic sequence from a standard part of the genome. This technique used to identify eight medicinal plants collected from the Ardabil province. DNA extraction was performed by modified CTAB method and PCR was performed with primers designed based on rbcL, trnH-psbA, matK Chloroplast barcodes and ITS nuclear barcode. Then, PCR products purified and sequenced. The percentage of amplification and sequencing success were assumed in samples respectively, 87 and 62, 75 and 37, 62 and 12, 75 and 37. The sequences were blasted with samples existed in NCBI database and Bioinformatics analyses were performed. In phylogenetic tree, the species belonging to the same genus were separated from other genus based on rbcL and trnH-psbA barcode sequences. Also, in ITS barcode only G. glabra organized with plants from same genus. In this study, barcoding of L. ledebourii with rbcL was done for the first time. SNPs were counted for barcodes of rbcL (less than 30), trnH-psbA (less than100), ITS (more than 200) and matK (less than 20). Thus, rbcL barcode due to high separation ability, low number of SNPs and universality in most species, was introduced as the best barcode. However, trnH-psbA and ITS barcodes due to related problem with direct sequencing of PCR products and lack of access to high quality sequences were identified as complementary barcodes. MatK barcode is not recommended for these samples because of the low ability of amplification and sequencing.

کلیدواژه‌ها [English]

  • Bracodes rbcL
  • trnH-psbA
  • matK and ITS
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