Genotypes-Independent Optimization of Nitrogen Supply for Isolated Microspore Cultures in Barley.

To establish a high-efficiency system of isolated microspore culture for different barley genotypes, we investigated the effects of nitrogen sources and concentrations on callus induction and plant regeneration in different barley genotypes. The results showed that the organic nitrogen sources greatly increased the callus induction, and the great reduction of total nitrogen sources would significantly decrease the callus induction. And the further optimization experiments revealed that the increasing of organic nitrogen sources was much important in callus induction while it seemed different in plant regeneration. Based on the great effects of organic nitrogen on callus induction, the medium of N6-ANO1/4-2000 might be the best choice for the microspore culture system. In addition, the phylogenetic analysis indicated that there were clear differences of genetic backgrounds among these barley genotypes, and it also suggested that this medium for microspore culture had widespread utilization in different barley genotypes.

Transcriptome analysis reveals translational regulation in barley microspore-derived embryogenic callus under salt stress.

KEY MESSAGE: Transcriptome analysis of barley embryogenic callus from isolated microspore culture under salt stress uncovered a role of translation inhibition and selective activation of stress-specific proteins in cellular defense. Soil salinity is one of the major abiotic stresses which constrains the plant growth and reduces the productivity of field crops. In this study, it was observed that the salt stress in barley isolated microspore culture impacted not only on the quantity of embryogenic callus but also on the quality for later differentiation. The barley microspore-derived embryogenic callus, a transient intermediate form linked cells and plants, was employed for a global transcriptome analysis by RNA sequencing to provide new insights into the cellular adaptation or acclimation to stress. A total of 596 differentially expressed genes (DEGs) were identified, in which 123 DEGs were up-regulated and 473 DEGs were down-regulated in the embryogenic callus produced from microspore culture under salt stress as compared to the control conditions. KEGG pathway analysis identified 'translation' (27 DEGs; 12.56 %) as the largest group and followed by 'folding, sorting and degradation' (25 DEGs; 11.63 %) in 215 mapped metabolic pathways. The results of RNA-Seq data and quantitative real-time polymerase chain reaction validation showed that the genes related to translation regulation (such as eIF1A, RPLP0, RPLP2, VARS) were down-regulated to control general protein synthesis, and the genes related to endoplasmic reticulum stress response (such as small heat shock protein genes) were selectively up-regulated against protein denaturing during microspore embryogenesis under continuous salt stress. These transcriptional remodeling might affect the essential protein synthesis for the cell development to fulfill totipotency under salt stress.