Optimization of DNA Isolation Protocol from Emerging Leaves of Black Pepper [Piper nigrum (L.)

Abstract

Black pepper (Piper nigrum L.) is a major spice crop and exported worldwide. The extraction of DNA from leaves of black pepper is challenging due to the presence of a significant amount of polysaccharides, polyphenolic compounds and other chemical constituents. Hence, the present study was aimed to standardize and optimize the DNA isolation protocol from the young folded leaves of black pepper. The emerging folded leaves of seven pepper accessions were selected and an efficient DNA extraction protocol from freshly collected emerging leaves of pepper was developed with modified cetyl trimethyl ammonium bromide (CTAB) method, which is a collective strategy of 2-mercaptoethanol, PVP (poly vinyl pyrrolidone), 1% sodium metabisulfite and 5M NaCl, which was used for efficient removal of polyphenols and polysaccharides. This protocol was pertinent to isolate genomic DNA from emerging leaf tissues of pepper. The presence of DNA was checked by 0.8% agarose gel; results showed that thick intact single band of high molecular weight confirming good quality. The genomic DNA yielded from fresh emerging leaves of seven pepper accessions was ranged from 75 - 120µg/g.