Data-Mining of Barley to Identify Salt Stress Hub Genes, Gene Expression Analysis and Recombinant Plasmid Construction.

Abstract

Salinity is one of the major abiotic stresses that limit the production and yields of agricultural crops worldwide. In order to identify key barley genes under salinity stress, the available metadata were examined by two methods of Cytoscape and R software. Next, the hub expression of the selected gene was evaluated under different salinity stress treatments and finally, this gene was cloned into cloning and expression vector and recombinant plasmid was made. In this study, we extracted salinity stress tolerant genes from several kinds of literature and also microarray data related to barley under salinity conditions from various datasets. The list of genes related to literature analyzed using string and Cytoscape. The genes from the datasets were first filtered and then the hub genes were identified by Cytoscape and R methods. Next, these hub genes were analyzed for the promoter. Ten hub genes were selected and their promoters were analyzed, the cis-element of which was often cis-acting regulatory element involved in the methyl jasmonate -responsiveness, common cis-acting element in promoter and enhancer regions and MYBHv1 binding site. Finally, the sedoheptulose-1,7-bisphosp gene (SBPase), which had the highest interaction in both gene lists and both types of gene networks, was selected as hub gene. Next, the expression of SBPase gene was examined in two variety of Youssef variety (salt tolerant) and Fajr variety (salt sensitive) under salinity stress (NaCl 100mM) at 0 (control), 3, 6, 12 and 24 hours after stress. The results showed that the expression of this gene increased with increasing the duration of stress in both varieties. Comparison of the two varieties showed that the expression of SBPase gene in the tolerant genotype was twice as high as sensitive. Finally, SBPase gene as a key gene for salinity stress was cloned in both cloning (pTG19) and expression (pBI121) vectors. According to our results, SBPase gene increased growth and photosynthesis in barley under various abiotic stresses, therefore, over-expression of this gene in barley is recommended to produce plants resistant to abiotic stresses.