Exogenous calcium application enhances salt tolerance of sweet sorghum seedlings

The intensification of global soil salinization has seriously decreased crop yield. At the same time, the rapid increase in population has raised the demand for crop yields. Reasonable use of saline-alkali land and cultivation of salt-tolerant crops through transgenic technology becomes more important. Sweet sorghum [Sorghum bicolor (L.)], as an important energy crop, resists abiotic stress effectively. In our previous salt tolerance gene-screening experiment of sweet sorghum, it was found that applying calcium ions could alleviate salt stress of sweet sorghum to some extent. Therefore, 10 and 20 mM CaCl2 were applied to explore the effect of exogenous calcium application on salt stress. Compared with the sweet sorghum treated by salt stress, the dry and fresh weight of the plants were significantly increased, and the content of malondialdehyde (MDA) was decreased after applying calcium ions. At the same time, diaminobenzidine and nitro blue tetrazolium chloride monohydrate staining results showed that the contents of H2O2 and O-2(-center dot) in the plants after calcium application were lower than those only treated with salt stress. To investigate the regulation mechanism of exogenous calcium application alleviating salt stress, RNA-seq analysis was performed using libraries established in Roma roots under normal and NaCl treatment. Combined with Gene Ontology and Kyoto Encyclopaedia of Genes and Genomes analysis, we found that calcium-related genes were more involved in the oxidation-reduction process and carbohydrate metabolism pathway. Among all differentially expressed genes, 12 known functional genes were further explored based on functional annotations and existing studies. To detect whether these 12 genes are related to salt response, we treated the Arabidopsis mutants of their homologous genes with 150 mM NaCl for 7 days. Among them, atglr3.5, atgad4, atiqm4, atglr2.8 and annat7 showed worse growth, lower survival rate and higher MDA content than WT under salt stress. This indicates that applying exogenous calcium can affect the expression of SbGR3.4, SbGADX1, SbIQM4, SbGR2.8 and SbAnnD7, which can regulate the expression of genes related to the lignin degradation, absorption of NH4+, and ubiquitination-proteasome pathway, energy metabolism and cleaning of ROS accumulation in sweet sorghum under salt stress, so as to enhance the salt tolerance of plants.