Standardization of DNA isolation and PCR parameters in Garcinia spp. for RAPD analysis

Abstract

Doyle and Doyle protocol with a little modification was used for the isolation of DNA from leaves and fruit rind of Garcinia spp. In case of leaves and dry fruit rind, the best DNA isolation was possible with 4% CTAB (100 mM Tris, 30 mM EDTA & 1.4 M NaCl), followed by 1.5% PVP and 0.3% mercaptoethanol. In case of fresh fruit rind, the DNA was best isolated with 2% CTAB (100 mM Tris, 30 mM EDTA & 1.4 M NaCl). PCR parameters were optimized using random RAPD primers. Other parameters included the DNA concentration at 50 ng/reaction, annealing temperature of 43°C, 2.5 mM dNTP in presence of 1 U Taq DNA polymerase and 2.0 mM MgCl2. The protocol will be of great help to study the genetic diversity of Garcinia germplasms.