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An improved protocol for genomic DNA extraction and purification from leaves of Terminalia Arjuna (Roxb.) Wight and Arnis

Author Affiliations

  • 1Tropical Forest Research Institute, Jabalpur-483001, Madhya Pradesh, India

Int. Res. J. Biological Sci., Volume 7, Issue (10), Pages 10-16, October,10 (2018)

Abstract

Several available protocols fail to produce good quality genomic DNA from leaves of Terminalia arjuna that contain the exceptionally high amount of interfering secondary metabolites and polysaccharides. To address this issue, a modified DNA isolation protocol was developed for obtaining quality genomic DNA. Briefly, leaf tissue was macerated in lysis buffer supplemented with PVP, sorbitol and sodium chloride. The macerated material was washed in sorbitol buffer for easy extraction and getting rid of mucilaginous substances present in the sample. Subsequently, the inclusion of high concentrations of B-mercaptoethanol, NaCl, and Triton-x in Cetyltrimethylammonium bromide (CTAB) removed polyphenols and polysaccharides. Phenol: Chloroform: Isoamyl alcohol washings took after by precipitation with NaCl proficiently expelled high protein and polysaccharide impurity. The purity of the isolated genomic DNA was checked spectrophotometrically and confirmed by the running of isolated DNA on 0.8% agarose gel electrophoresis. The yield of pure genomic DNA ranged from 120 to 570ng/µl and purity index (A260/A280) ratio from 1.65 to 2.0. The isolated genomic DNA optimized the RAPD analysis and showed clear amplification in all the germplasm tested.

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