BackgroundBasil is one of the most famous herbs, which has broad usage as a fresh vegetable and therapeutic and pharmaceutical services. The main abiotic stress limiting basil production globally is drought. As a result, appropriate drought screening-which effectively separates high-yielding but drought-sensitive genotypes from drought-tolerant genotypes-is necessary for the optimal selection of high-yielding basil cultivars under drought stress conditions. So, a split plot experiment with three replications based on a completely randomized design were carried out in a pot under field conditions for this investigation. Water levels (full irrigation or control, moderate stress, and severe stress) were assigned as main plots, while 22 basil accessions were given as sub-plots. In this study, leaf yield as well as physio-biochemical traits had measured on accessions.ResultsOur results revealed large variation in yield, essential oil (%), protein, proline, chlorophyll, total phenol and flavonoids traits across the 22 accessions. The percentage of leaf yield reduction in moderate drought stress than normal conditions showed that G1 (−6.5%), G17 (−7.05%), G20 (−9.01%), and G12 (−10.9%) accessions had the least changes, respectively. Although in severe drought stress than normal conditions, the G1 (−32.01%), G12 (−33.12%), G4 (−33.24%), G7 (−34.11%), and G17 (−34.93%) accessions had the least amount of change in plant leaf yield, respectively. Furthermore, the highest yield reduction occurred in moderate and severe stress conditions in G18 (−25.36%) and G8 (−42.98%) accessions, respectively. Cluster analysis based on the ward method in both conditions (moderate and severe drought conditions) placed the accessions in three groups, and accessions were identified as tolerant, whose average traits in that group were higher than the total average. The principal component analysis also showed that in moderate drought conditions, the first two components explained about 95.28% of the total variation, while in severe drought conditions, these two components explained about 96.37% of the total variation.ConclusionsThe different multivariate analyses (cluster analysis, PCA, mean comparison) were used to identify tolerant and sensitive accessions based on all traits. The accessions G3, G4, G6, and G7 were found to be tolerant to stress, while G10, G15, G16, and G20 were found to be sensitive to drought. These accessions are a useful step in producing drought-tolerant, high-yielding accessions and can be utilized in breeding programs for basil.