Successfully promoting drought tolerance in wheat genotypes will require several procedures, such as field experimentations, measuring relevant traits, using analysis tools of high precision and efficiency, and taking a complementary approach that combines analyses of phenotyping and genotyping at once. The aim of this study is to assess the genetic diversity of 60 genotypes using SSR (simple sequence repeat) markers collected from several regions of the world and select 13 of them as more genetically diverse to be re-evaluated under field conditions to study drought stress by estimating 30 agro-physio-biochemical traits. Genetic parameters and multivariate analysis were used to compare genotype traits and identify which traits are increasingly efficient at detecting wheat genotypes of drought tolerance. Hierarchical cluster (HC) analysis of SSR markers divided the genotypes into five main categories of drought tolerance: four high tolerant (HT), eight tolerant (T), nine moderate tolerant (MT), six sensitive (S), and 33 high sensitive (HS). Six traits exhibit a combination of high heritability (>60%) and genetic gain (>20%). Analyses of principal components and stepwise multiple linear regression together identified nine traits (grain yield, flag leaf area, stomatal conductance, plant height, relative turgidity, glycine betaine, polyphenol oxidase, chlorophyll content, and grain-filling duration) as a screening tool that effectively detects the variation among the 13 genotypes used. HC analysis of the nine traits divided genotypes into three main categories: T, MT, and S, representing three, five, and five genotypes, respectively, and were completely identical in linear discriminant analysis. But in the case of SSR markers, they were classified into three main categories: T, MT, and S, representing five, three, and five genotypes, respectively, which are both significantly correlated as per the Mantel test. The SSR markers were associated with nine traits, which are considered an assistance tool in the selection process for drought tolerance. So, this study is useful and has successfully detected several agro-physio-biochemical traits, associated SSR markers, and some drought-tolerant genotypes, coupled with our knowledge of the phenotypic and genotypic basis of wheat genotypes.