دستورالعمل ساده و کارآمد استخراج DNA برای برگ‌های قدیمی و هرباریومی تمشکین (تیره مارچوبه‌ایان)

نویسندگان

1 گروه زیست شناسی، واحد مشهد، دانشگاه آزاد اسلامی، مشهد، ایران

2 گروه زیست شناسی، واحد مشهد، دانشگاه آزاد اسلامی، مشهد، ایران؛ گروه زیست شناسی، دانشکده علوم، دانشگاه فردوسی مشهد

3 دانشگاه آزاد اسلامی مشهد

چکیده
DNA استخراج شده با کیفیت بالا در بدست آمدن باندهای واضح در مرحله الکتروفورز و نتایج صحیح در تعیین توالی نقش مهمی ایفا می­کند. معمولا CTAB یکی از روش های رایج برای استخراج DNA است که برای برگ­های قدیمی نمونه­های هرباریومی سرده تمشکین (Bellevalia) که متابولیت های ثانویه با اثر بازدارندگی دارد مناسب نسیت. برای حل این مسئله، تغییراتی در مراحل مختلف CTAB اعمال شد و ترکیبات زغال فعال، فنل، استات پتاسیم و RNase استفاده شد و آب دیونیزه جایگزین Tris EDTA شد تا ناخالصی­ها حذف شده و DNA با کیفیت تری بدست آید. داده­های میزان جذب DNA روش جدیدبا روش CTAB در طول موج­های ۲۶۰ و ۲۸۰ نانومتر مقایسه شد. نتایج، تغییر ۱۵تا۸۰ برابری در غلظت DNA را نشان داد. این مطالعه روش اصلاح شده­ای را پیشنهاد می­کند که امکان استخراج DNA با کیفیت بالاتری نسبت به روش CTAB را از برگ­های قدیمی نمونه های هرباریومی Bellevalia امکان­پذیر می­کند.






کلیدواژه‌ها


عنوان مقاله English

A simple and efficient DNA extraction protocol for old herbarium leaves of Bellevalia (Asparagaceae, Scilloideae)

نویسندگان English

Lida Jalali Roudsary 1
Azarnoosh Jafari 1
Jamil Vaezi 2
Ehsan Karimi 3
1 Department of Biology, Mashhad Branch, Islamic Azad University, Mashhad, Iran
2 Department of Biology, Mashhad Branch, Islamic Azad University, Mashhad, Iran; Department of Biology, Faculty of Sciences, Ferdowsi University of Mashhad, Mashhad, Iran
3 Department of Biology, Mashhad Branch, Islamic Azad University, Mashhad, Iran
چکیده English

High-quality DNA extraction plays an important role to make sharp bands in the gel electrophoresis and also produces clean chromatograms. Usually, DNA extract is delivered using the modified CTAB method but this method cannot obtain high-quality DNA for molecular analysis from old dried leaves of Bellevalia due to having different chemical compounds which inhibit to obtain a clear DNA extraction. To solve the problem, in different phases of a modified CTAB protocol, activated carbon, phenol, potassium acetate, RNase and deionized water (instead of TE) were applied to eliminate contaminants and improve obtained high-quality DNA. The DNA absorption data from the modified method were compared with CTAB method in 260 and 280 nm wavelengths. The results showed a 15-80-fold change in the DNA concentration. This study suggests the modified method of extraction is more efficient in the quality of DNA obtained from the little amount of herbarium old leaves of Bellevalia than that of the CTAB method.


کلیدواژه‌ها English

activated charcoal
Bellevalia
DNA purification
Phenol
nanodrop
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دوره 9، شماره 2 - شماره پیاپی 32
تابستان 1401
صفحه 124-131

  • تاریخ دریافت 18 خرداد 1405
  • تاریخ انتشار 18 خرداد 1405