Abstract Biochar (BC) and plant growth-promoting microbes (PGPR) could represent a suitable agronomical strategy to mitigate the impacts of drought in arid agro-environmental conditions. However, there is currently little understanding of the synergistic benefit of combining BC and PGPR to increase drought tolerance in oilseeds. In this study, the physiological response of two water-stressed canola ( Brassica napus L.) plants subjected to the application of BC obtained from waste wood of Morus alba applied solely or in combination with PGPR strains ( Pseudomonas sp.) was evaluated. The experiment consists of two genotypes and nine treatments [(C-Control, T1-15 days drought (15DD), T2-30 days drought (30DD), T3-15 days of drought + PG (15DD + PG), T4-30 days of drought + PG (30DD + PG), T5-15 days drought + biochar (15DD + BC), T6-30 days drought + biochar (30DD + BC), T7-15 days drought + biochar + PG (15DD + BC + PG), T8-30 days drought + biochar + PG (30DD + BC + PG)]. Drought stress decreased emergence energy (EE), leaf area index (LAI), leaf area ratio (LAR), root shoot ratio (RSR), moisture content of leaves (MCL), percent moisture content (%MC), moisture content of shoot (MCS) and moisture content of root (MCR), and relative water content (RWC) in both varieties of Brassica napus L., which in contrast, it is increased by the collective application of both biochar and PGPR. In both varieties, N, P, K, Mg, and Ca concentrations were highest in all the biochar and PGPRs separate and combined treatments, while lowest in 15 and 30 days drought treatments. Osmolyte contents like Glycine betaine (GB) and sugar remarkably increased in the stress condition and then reduced due to the synergistic application of biochar and PGPR. Drought stress has a repressive effect on the antioxidant enzymatic system like Peroxidase (POD), Superoxide dismutase (SOD), and glutathione reductase (GR) as well as total flavonoids, phenolics, and protein content. The antioxidant enzymes and phenolic compounds were dramatically increased by the combined action of biochar and PGPRs. A significant increase in EE, LAR, RSR, and RWC under 15 and 30 days drought conditions, evidently highlighting the synergistic effect of BC and PGPR. The results conclude a substantial and positive effect of the combined use of BC and PGPR strains on canola's response to induced drought stress, by regulating the physiological, biochemical, and agronomic traits of the plants. Graphical Abstract