PCR Optimization Based on Inter-Primer Binding Site (iPBS) Markers for Genetic Diversity Analysis in Durum Wheat Genotypes

In this study, PCR optimization was performed using iPBS (inter-primer binding site) molecular markers to determine the genetic diversity of durum wheat genotypes. Optimizing PCR conditions to enhance the efficiency of iPBS markers was a critical step. To improve the accuracy of the molecular analysis, gradient testing was conducted with 20 different iPBS primers to determine the optimum annealing temperatures. The results showed that 11 primers produced polymorphic bands, and the temperatures at which these primers bound most efficiently were identified. Correctly determining the annealing temperatures of the primers directly impacted the success of the PCR process, ensuring reliable DNA amplification.