Abstract

Extraction of DNA in many plants is difficult because of the presence of metabolites that interfere with DNA isolation procedures and downstream applications such as DNA restriction, replications, amplification, as well as cloning. Modified procedure based on the hexadecyltrimethyl ammoniumbromide (CTAB) method is used to isolate DNA from tissues containing high levels of polysaccharides. The procedure is applicable to both ripped and unripe fruits of Annona senegalensis. This modified CTAB (2%) protocol include the use of 1.4 M NaCl, 1% polyvinylpyrrolidine (PVP), 1% mercaptoethanol and 100% absolute ethanol in the extraction as well as reducing the centrifugation times during the separation and precipitation of the DNA. This method solved the problems of DNA contamination, degradation and low yield due to binding or co-precipitation with starches. The isolated DNA proved amenable to polymerase chain reaction (PCR) amplification and restriction digestion. This technique is fast, reproducible, and can be applied for simple sequence repeats (SSR)-PCR markers identification.

Key words: Annona senegalensis, genomic DNA, fruits, modified, markers.

Abbreviation

Abbreviations: PCR, Polymerase chain reaction; RFLPs, restriction fragment length polymorphisms; RAPD, randomly amplified polymorphic DNA; SSR, simple sequence repeats; SDS, sodium dodecyl sulphate; CTAB, cetyl-methyl ammonium bromide; AFLPs, amplified fragment length polymorphism; ISSR, inter simple sequence repeat; PVP, polyvinylpyrrolidine.