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DNA markers help detect the intraspecific genetic diversity of yeast strains. Eight ISSR (Inter Simple Sequence Repeats) primers were used to assess the intraspecific diversity of <i>Saccharomyces cerevisiae</i> (<i>n</i> = 96) from different populations (<i>n</i> = 3), evaluate the technological characteristics, and investigate trait-loci associations. The primers amplified 154 reproducible and scorable bands, of which 79.87% were polymorphic. The UPGMA (unweighted pair group method with arithmetic mean) dendrogram clustered 96 isolates into two main clusters, supported by STRUCTURE HARVESTER results (Δ<i>K</i> = 2). Analysis of molecular variance (AMOVA) indicated significant genetic differences between (15%) and within the populations (85%) (<i>p</i> < 0.001). Twenty-nine genetically distinct strains were selected for the technological characterization. Principal component analysis (PCA) revealed that five strains with high fermentation capacity, leavening activity, high growth index at 37 °C, and harsh growth conditions were technologically relevant. Trait-loci association analyses indicated that the highest correlation (<i>r</i> = 0.60) was recorded for the fermentation capacity on the 8th and 113th loci, amplified by ISSR-1 and ISSR-6 primers, respectively (<i>p</i> < 0.05). The strains yielding high performances and the associated loci amplified by ISSR markers possess a high potential to generate locus-specific primers to target the strains with high fermentation capacity.