A modified DNA extraction protocol and its utility in seed genetic purity assessment

Abstract

The isolation of high quality DNA in sufficient quantity is the basic need in all PCR-based techniques. None of the currently available methods provides a risk-free, time and cost-effective DNA extraction protocol which is essentially required when a large number of plant/seed samples are analyzed in marker-assisted techniques. In the present study, we describe a simple, inexpensive and reliable method for isolating DNA from six plant species (Glycine max, Gossypium hirsutum, Sesamum indicum, Vigna radiate, Vigna Aconitifolia and Vigna unguiculata), avoiding costly and risky steps such as, use of liquid nitrogen, lyophilization and dehydration. The isolated DNA (Mw∼40 kb) proved amenable in PCR amplification, restriction digestion and cloning, and comparable to CTAB and potassium acetate methods in quantity and quality. Also, the utility of this method was demonstrated for RAPD-based genetic purity assessment of cotton hybrid through DNA sampling following two-dimensional growth strategy and the results were found comparable with the grow-out test irrespective of the matrix size. We recommend the use of this method for all PCR-based techniques such as, genetic purity assessment and rapid genotyping in marker-assisted breeding programmes.