Protocol optimization for deoxyribonucleic acid (DNA) extraction from dried, fresh leaves, and seeds of groundnut (Arachis hypogaea L.)

Abstract

Consistent isolation of best quality deoxyribonucleic acid (DNA) from peanut (Arachis hypogaea L.) is particularly problematic due to the presence of phenolic compounds and polysaccharides. Inconsistencies in extraction results can be attributed to the age and growth stages of the plant material analyzed. Mature leaves have higher quantities of polyphenols, tannins and polysaccharides that can contaminate DNA during isolation. In this study, we used fresh and dried leaves as well as seeds for optimization of high quality DNA isolation protocols from A. hypogaea. The DNA extracted with three different methods cetyltrimethylammonium bromide (CTAB), sodium dodecyl sulfate (SDS), and cesium chloride (CsCl) density gradient) were comparatively studied by polymerase chain reaction (PCR) analysis in terms of quantity and quality. High quality genomic DNA was obtained from fresh leaves by modified CTAB methods. The DNA obtained ranged from 1 to 2.5 ng/μl. DNA obtained by this method was strong and reliable showing its compatibility for simple sequence repeat (SSR) analyses. The SDS based methodology give large quantities of DNA contaminated with polysaccharides. Fresh leaves also gave best result in SDS method. The quantity and quality of DNA obtained was very poor in all the tested methods in case of dried leaf tissues. The current protocol will probably be useful for the extraction of high-molecular weight DNA from other plant materials containing large amounts of secondary metabolites and essential oils.

Key words: Polysaccharides, polyphenols, tannins, cetyltrimethylammonium bromide (CTAB), sodium dodecyl sulfate (SDS), cesium chloride (CsCl), secondary metabolites, SSR.