Extraction of high-quality genomic DNA from different plant orders applying a modified CTAB-based method

Nadia Aboul-Ftooh Aboul-Maaty,Hanaa Abdel-Sadek Oraby

doi:10.1186/s42269-019-0066-1

Nadia Aboul-Ftooh Aboul-Maaty, Hanaa Abdel-Sadek Oraby

Open Access

https://doi.org/10.1186/s42269-019-0066-1

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Abstract

BackgroundReliable measurement of DNA concentration and purity is important for almost all molecular genetics studies. Different plant species have varying levels of polysaccharides, polyphenols, and other secondary metabolites which combine with nucleic acids during DNA isolation and further affect the quality of the extracted DNA. The current extraction protocol is based upon the conventional cetyl trimethylammonium bromide (CTAB) method with further modifications for the extraction of DNA from variable plant seeds and crops belonging to seven different orders. The principle modifications currently employed for DNA extraction involved the use of higher CTAB concentration and higher levels of 2-β-mercaptoethanol. Additionally, higher concentrations of sodium chloride and potassium acetate were added simultaneously with absolute ice cold isopropanol for the precipitation of DNA free from polysaccharides.Results and conclusionThe prescribed modifications in the present method establish a quick and efficient standardized protocol for DNA extraction from different plant orders. The current extraction protocol, therefore, can be of great value for molecular analysis involving large numbers of different plant samples from different orders. These modifications consistently produced pure and high-quality DNA suitable for further molecular analysis. Successful PCR amplification with random amplified polymorphic DNA primer, NPTII gene, and the complete digestion of the isolated DNA with the HindIII restriction enzyme validated the quality of the isolated DNA. Moreover, it reflects the efficiency of the protocol and proves its suitability for further applications for the assessment of food safety, detection of genetically modified (GM) crops, and conservation of biodiversity.

Highlights

Reliable measurement of Deoxyribonucleic acid (DNA) concentration and purity is important for almost all molecular genetics studies
Reliable measurement of DNA concentration and purity is important for the assessment of food safety, especially with the increase of the global cultivation area of genetically modified (GM) crops (Ateş Sönmezoğlu and Keskin 2015)
Applying the present standardized method, the extracted DNA concentrations varied with the different plant species used in the present work (Table 2)

Summary

Introduction

Reliable measurement of DNA concentration and purity is important for almost all molecular genetics studies. Different plant species have varying levels of polysaccharides, polyphenols, and other secondary metabolites which combine with nucleic acids during DNA isolation and further affect the quality of the extracted DNA. Reliable measurement of DNA concentration and purity is important for the assessment of food safety, especially with the increase of the global cultivation area of genetically modified (GM) crops (Ateş Sönmezoğlu and Keskin 2015). Many difficulties have been reported for isolating good-quality DNA from plants (Novaes et al 2009; Silva 2010; Moreira and Oliveira 2011) These difficulties were attributed to the fact that different plant species have varying levels of polysaccharides, polyphenols, and other

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Discussion

Conclusion