The seeds were surface sterilized and then soaked for two hours and sown in soil mixture having 3 parts of garden soil, 1 part of sand and 1 part of manure. The experiment was carried out in earthen pots of uniform size each containing 5 Kg of the soil mixture. Before sowing, the soil was amended with K₂CrO₄ for Cr treatments (0 µM/Kg), and Na₂SeO₄ for Se treatments (0 µM/Kg), both alone and in combinations. The concentration used for Cr was 50% inhibitory concentration (IC50), while for Se, the most stimulatory concentrations as observed from preliminary studies were used. The pots were kept in natural environmental conditions and were watered regularly. The experiment was conducted in triplicates. The harvesting of the plants was done after 30 days of sowing and stored at -20 ℃ . Some harvested plants were also dried by keeping them in hot air oven for 24 h at 80 ℃.

Se application aided in improving plant growth, reducing the oxidative damage and strengthening the antioxidative defence system in plants raised in soils with binary combinations of Cr and Se. Photosynthesis, which is one of the vital physiological processes, was positively influenced with application of Se. It helped in minimising the toxicity of Cr and enhanced the contents of pigments. The efficiency of photosynthetic machinery was further strengthened by the increase of net photosynthetic rate, transpiration rate, stomatal conductance and intercellular CO₂ concentration, and hence indicated its importance in combating stress. The study also highlighted the role of Se in enhancing the contents of secondary metabolites which play an important role in heavy metal chelation, complex formation and ROS scavenging, thereby reducing the chances of Cr to cause physiological damage.

A significant modulation in gene expression was observed in B. juncea in response to Cr and Se. The gene responsible for H₂O₂ production is respiratory burst oxidase (RBO) which showed a significant upregulation in its expression by 3.63 folds in response to Cr treatment. Se at 2 µM/Kg in combination with 300 µM/Kg Cr caused decrease by 1.62 folds in the expression of RBO gene.An increase in expression was observed SOD, CAT and GST-1 by 2.75 folds, 2.82 folds and 2.03 folds respectively in response to Cr. However, Cr treatment resulted in a reduction of relative expression of POD and GR genes by 0.54 and 0.61 folds respectively in leaves of B. juncea plants. The combined treatment of Se and Cr aided in reducing Cr toxicity by increasing the expression of genes coding all these enzymes. Maximum increase in expression in case of CAT (4.68 folds), GR (2.08 folds) and GST-1 (2.98 folds) was observed at binary combination of 4 µM/Kg Se and 300 µM/Kg Cr. For SOD, 4.25 folds increase in gene expression was observed at 6 µM/Kg Se and 300 µM/Kg Cr. The expression of POD enhanced by 1.75 folds at the concentration of 2 µM/Kg Se and 300 µM/Kg Cr. The genes coding forcholrophyllase (CHLASE) chalcone synthase (CHS) and phenylalanine ammonialyase (PAL) showed enhanced expression of 2.47 folds, 1.79 folds and 2.07 folds respectively in the plants raised in Cr spiked soils. The co-application of Se and Cr helped in increasing the expression of CHS and PAL, while aided in reducing the expression of CHLASE. The concentration of 4 µM/Kg for Se proved to be most beneficial for enhancing the gene expression of PAL by 3.92 folds, while the same concentration caused a decline in the expression of CHALSE by 1.65 folds. However, for CHS expression, 6 µM/Kg Se caused an increase by 2.52 folds. Statistical analysis by one-way ANOVA and MLR supported the observations. The values of beta-regression coefficients for Se indicated the stress alleviating effects of Se for all the genes.