Drought stress at germination stage is an important environmental stress limiting crop yield. Hence, our study investigated comparative root transcriptome profiles of four contrasting soybean genotypes viz., drought-tolerant (PI342618B/DTP & A214/DTL) and drought-sensitive (NN86-4/DSP & A195/DSL) under drought stress using RNA-Seq approach. Total of 4850 and 6272 differentially expressed genes (DEGs) were identified in tolerant (DTP & DTL) and sensitive (DSP & DSL) genotypes, respectively. Principle component analysis (PCA) and correlation analysis revealed higher correlation of DTP with DTL. Both gene ontology (GO) and MapMan analyses showed drought response was enriched in the DEGs associated with water and auxin transport, cell wall/membrane, antioxidant activity, catalytic activity, secondary metabolism, signaling and transcription factor (TF) activities. Out of 981 DEGs screened from above terms, only 547 showed consistent opposite expression between contrasting genotypes. Twenty-eight DEGs of 547 were located on Chr.08 rich in QTLs and “Hotspot regions” associated with drought stress, and eight of them showed non-synonymous SNP polymorphism. Hence, ten genes (including above eight genes plus two hub genes) were predicated as possible candidates regulating drought tolerance, which needs further functional validation. Overall, the transcriptome profiling provided in-depth understanding about the genetic mechanism and candidate genes underlying drought tolerance in soybean.