*3.7. Priming Effect of SPM on the Production of Endogenous ABA, JA and Free SA Levels under Cr Stress*

The ABA, JA and SA levels were examined in seeds primed with water or SPM, with and without Cr stress. The exposure of Cr alone led to elevated contents of ABA (2.83 and 2.49-fold), JA (3.32 and 3.29-fold) and SA (2.48 and 1.89-fold) in both varieties CY927 and YLY689, individually, when compared to controls (Figure 5a–c). Under Cr stress, seed priming with SPM applications significantly decreased the ABA level (Figure 5a), whereas no significant difference was observed in the levels of JA under Cr treatments (Figure 5b). Nevertheless, the level of SA was enhanced significantly (2.23 and 1.76-fold) in both rice cultivars (CY927 and YLY689), individually, by the applications of seed primed SPM under Cr stress (Figure 5c). A similar pattern was seen in the seedlings of both rice cultivars, with YLY689 displaying more obvious effects than CY927.

**Figure 4.** Effects of SPM on the activities of (**a**) superoxide dismutase (SOD) in shoots, (**b**) superoxide dismutase (SOD) in roots, (**c**) ascorbate peroxidase (APX) in shoots, (**d**) ascorbate peroxidase (APX) in roots, (**e**) peroxidase (POD) in shoots, (**f**) peroxidase (POD) in roots, (**g**) catalase (CAT) in shoots, and (**h**) catalase (CAT) in roots of both rice varieties (CY927 and YLY689) under chromium (Cr) stress. Values are mean ± SE of three independent replicates and different letters (a–f) above bars show a significant difference between treatments by LSD test at *p* < 0.05.

**Figure 5.** Effects of SPM on the endogenous production of phytohormones including (**a**) abscisic acid (ABA), (**b**) Jasmonic acid (JA), (**c**) salicylic acid (SA) and expressions of phytohormones-associated genes (**d**) *PR1*, (**e**) *PR2*, and (**f**) *NPR1* in two rice cultivars (CY927 and YLY689) under chromium (Cr) stress. Values are mean ± SE of three independent replicates and different letters (a–f) above bars show a significant difference between treatments by LSD test at *p* < 0.05.

#### *3.8. Priming Effect of SPM on Hormone-Associated Gene Expression Analysis under Cr Stress*

Under Cr stress, the transcript levels of *PR1*, and *PR2* genes were subsequently upregulated in both CY927 and YLY689 compared to relative controls. Furthermore, the transcription levels of both these genes were markedly stimulated within seeds primed by SPM against Cr-induced stress as compared to the treatments of Cr alone in both rice cultivars (Figure 5d,e). The expression levels were noticeably upregulated in CY927 as compared to YLY689 (*p* < 0.01). Likewise, the upregulation in *NPR1* levels was noticed under Cr stress in both rice cultivars. Nevertheless, their expression levels were further increased in the seeds treated with SPM priming under Cr exposure (Figure 5f). An identical drift was noted in both rice varieties. The upregulation was more pronounced in the expression of the *NPR1* gene as compared to *PR1*, and *PR2* genes. This indicated that *NPR1* has a dynamic role in stimulating the phytohormone synthesis and mitigation of Cr toxicity. In addition, gene expression analysis supported the notion that SPM mediated modifications in SA contents under Cr applications. Our findings clearly represented that SPM has extensive participation in rice tolerance against Cr stress and modulates the levels of transcription of certain stress-responsive genes associated with phytohormones.
