Assessment of Various Techniques for Extracting Genomic DNA from Herbarium Specimens of Myosotis
Subject Areas : Plant Biologyzahra ghasemzadeh 1 , Maryam Khoshsokhan mozaffar 2
1 - Ms of Biology, Qo.C. Islamic Azad University, Qom, Iran
2 - Associate Prof. Department of Biology, Qo.C. Islamic Azad University, Qom, Iran
Keywords: DNA Extraction, CTAB, Myosotis, Dellaporta, Secondary Metabolites,
Abstract :
Introduction: DNA extraction is a crucial technique in molecular biology. However, extracting DNA from various plants can be challenging due to the presence of secondary metabolites. This study aims to discover an effective method for extracting DNA from Myosotis species, part of the borage family.
Materials and Methods: Three extraction methods were employed on five Myosotis species: unmodified CTAB, modified CTAB, and Dellaporta.
Results: The findings revealed that using unmodified CTAB resulted in a weak band only for M. stricta (collection 2007). In contrast, the modified CTAB method produced strong bands for M. stricta (collections 2007 and 2012) and M. asiatica (collection 2004), while M. sparsiflora (collection 1980) and M. sylvatica (collection 1988) yielded weaker bands. The Dellaporta method showed a band for M. asiatica (collection 2004).
Conclusion: The modifications in the CTAB method, which included increasing the concentration of 60% mercaptoethanol (to prevent polyphenol oxidation) and adding ammonium acetate (for improved separation of histones from nucleic acids), resulted in better DNA concentration. Given the potential for DNA degradation in older samples, the modified CTAB method provided superior quality and quantity of extracted DNA.
1. Doyle JJ, Doyle JL. Isolation of plant DNA from fresh tissue. Focus; 1990 12:13-15.
2. Dellaporta SL, Wood J, Hicks JB. A plant molecular DNA minipreparation version 2. Plant Mol Biol Rep. 1983;1(1):19-21.
3. Kang TJ, Yang MS. Rapid and reliable extraction of genomic DNA from various wildtype and transgenic. Biotechnology; 2004 6750:4-20.
4. Khanuja SPS, Shasany AK, Darokar MP, Kumar S. Rapid isolation of DNA from dry and fresh samples of plants producing large amounts of secondary metabolites and essential oils. Plant Molecular Biology Reporters. 1999; 17: 1–7.
5. Khatamsaz, M, Flora of Iran, No. 39 – Boraginaceae, Forest and Rangeland Research Institute Publications; 2002.
6. Alidoosty-shahraky Z, Farrokh P, Amirahmadi A, Rezaei A, Ghasemian J. Comparison of five genomic DNA extraction methods in Iranian medicinal plant, Echium amoenum. J Med Plants. 2023;36(3):216-231.
7. FatemiFard SZ, Masoumiasl A. DNA extraction from dried specimens of plant and fungi in a fast, secure, and non-liquid nitrogen method. Iran J Plant Biotech. 2019;14(3):20-27.
8. Saboora A, Amiri Rad M, Asgarani E, Radjabian T. Comparison of three methods for genomic DNA extraction from fresh and herbarium leaf specimens of Achillea wilhelmsii. Nova Biol Res. 2019;5:458-465.
9. Zabiti SM, Ismaeili Maddah Arefi H, Nazarian Firouzabadi F, Mojiri F. Separation of secondary metabolites in DNA extraction process of thyme and analysis using GC-MS. Cell Mol Res. 2014;27(1):66-74.
10. Hoseinpour-Azad N, Nematzadeh GH. Introducing a fast and simple method for DNA extraction from Persian hogweed (Echium amoenum). Plant Ecosyst. 2012;8(31):111-120.
11. Banitalebi-Dehkordi S, Jafari-Sayyadi MH, Alavi-Samani SA. Optimization of CTAB method for DNA extraction from Iranian oak leaf tissue Quercus brantii Lindl. 8th National Biotechnology Congress of I.R. Iran & 4th National Conference on Biosecurity; 2013; Tehran, Iran.
12. Salehi K, Mozaffarian-Firoozabadi F, Esmaeili A, Zabeti SM. Determining the appropriate method for extracting genomic DNA from the leaves of medicinal plant chamomile. Nat Conf Med Plants. 2010.
