Optimization of DNA isolation and PCR- RAPD methods for molecular analysis of cluster bean (Cyamopsis tetragonaloba (L.) Taub) with cultivars of semiarid region

This paper present the optimization of DNA isolation and PCR conditions for RAPD analysis of Cyamposis tetragonaloba. The method involves the utilization of modified CTAB extraction by using higher concentration of NaCl and CTAB along with successive long-term chloroform: iso-amyl alcohol extractions. The yield of DNA ranged from 15–21 μg/gm of the leaf tissue and the purity (ratio) was between 1.5–1.7 indicating minimal levels of contaminating metabolites. RAPD protocol was optimized by using higher concentration of MgCl₂ (3 mM), lower concentrations of primer (10 μM) and Taq polymerase (0.5 units), 50 ng of template DNA and an annealing temperature 37°C which resulted optimal amplification. After implementing various modifications, it can be concluded that this protocol for DNA isolation and RAPD profiling is suitable to produce scorable and clear bands in cluster bean.