Abstract
The isolation of intact, high-molecular-mass genomic DNA is essential for many molecular biology applications including long range PCR, endonuclease restriction digestion, southern blot analysis, and genomic library construction. Many protocols are available for the extraction of DNA from plant material, but obtain it is difficult in many plants because of metabolites that interfere with DNA isolation procedures and subsequent applications. With frame of the present work, we developed the first reliable and efficient method for isolating Allium roseum L. genomic DNA that is free from polysaccharides and polyphenols. This protocol uses 100 mM Tris-HCl (pH 8), 20 mM EDTA, 1.4 M NaCl, 3% PVP (polyvinylpyrrolidone 40.000), 3% mercaptoethanol, and an incubation at 65 C for 1 h. The purity of isolated genomic DNA was confirmed by spectrophotometric analyses (A260/230 ratio of 1.947, A260/280 of 1.804). DNA was obtained in the amount of 189 mu g per gram of leaf material, and it proved amenable to restriction digestion.