Decreasing freshwater supply has led to the consideration of treated wastewater as an alternative source for
agricultural irrigation. However, the higher content of heavy metals restricts their use in agricultural purposes.
In the current study, a set of 181 SSR markers were used to perform association mapping in a structured sorghum
population consisting of 107 accessions evaluated under tap-water and heavy-metals-containing water irrigation
conditions. Significant differences between optimal and stressed growing conditions were shown in all evaluated
phenotypic traits. Association mapping revealed 14 significant associations between 12 SSR markers and heavy
metals stress indices of phenotypic traits (R2 = 11.54–30.85%). BLASTP annotation of QTLs genomic sequences
identified 102 gene homologs, of which 19 are known to be implicated in phytoremediation and heavy metals
tolerance. All nineteen genes exhibit differential expression patterns in heavy metals tolerant and susceptible
sorghum accessions, and their transcriptional levels were highly elevated under heavy metals stress, indicating a
possible functional association among these genes and provides strong evidence for their role in phytoremediation and heavy metal stress tolerance. Clustering and expression patterns of pentatricopeptide repeat and
zinc finger protein genes suggest crucial roles of those genes in phytoremediation and heavy metals tolerance.