Physiological Changes Of Salt Tolerance And Transcriptome Analysis Of Suaeda Glauca Under Salt Stress

With the aggravation of global soil salinization,the cultivation of various agricultural crops will face great challenges.Suaeda glauca(Bunge),as a typical halophyte,has strong salt-tolerance ability,which is of great value in food,medicine and environmental improvement.Through the detection of morphological changes and physiological level of S.glauca under salt stress,observation on the changes of salt treatment,and then select the appropriate concentration and time of salt treatment according to the guidance of physiological indicators.Through transcriptome sequencing,the related genes of salt tolerance of S.glauca were discovered.Seedlings of S.glauca were treated with 300 mmol/L Na Cl for 0,4,8,12,24 and48h respectively to observe their appearance during the process.The changes of cell structure and the surface of leaves were observed by paraffin section and scanning electron microscope.It was found that the plants lose turgor and wilted at 2h after salt stress,and began to adapt to the salt environment at 4h,and eventually recovered upright at 8h.The results of paraffin section showed that the parenchyma cells in the salt treated leaves were larger than those of control,which was of great significance for ion localization and osmotic regulation.It was found by SEM that the stomatal aperture and the proportion of stomata on the surface of leaves after salt treatment decreased obviously.The content of proline,soluble protein and soluble sugar in S.glauca was determined.The results showed that the content of proline and soluble protein increased with the increase of salt treatment time before 24h,and reached the maximum in 24h.The soluble sugar reached the maximum at 12h,and decreased after12h,but was still much larger than the control group.Furthermore,The Na⁺/K⁺ratio of S.glauca leaves was measured and was found to increase with the treatment time and reached the highest in 24h,and decreased in 48h.Further research showed that the chlorophyll content decreased with the treatment time,the stomatal conductance,transpiration rate and net photosynthetic rate of leaves in salt treatment group were lower than those in the control group,while the intercellular CO₂ concentration was higher than that in the control group.Malondialdehyde(MDA)content in antioxidant system increased with the treatment time,reached the highest value in 24h,and began to decline in 48h.The activities of Peroxidase(POD)and Catalase(CAT)increased with the salt treatment time.The activities of CAT enzyme reached the highest value in 24h,and the activities of POD enzyme reached the highest value in 12 h and then tended to be stable.In addition,the relationship between SOD activity and salt treatment time is not clear.A total of 30303564 reading sequences were obtained by transcriptome sequencing.Through the annotation of gene function,Gene Ontology(GO)was enriched into three categories:biological process,cell composition and molecular function,and a total of 72871 genes were annotated.Among them,Most of genes are mainly involved in the process of cell metabolism,cell regulation,cell composition and binding function.Through Eukaryotic Orthologous Groups of proteins(KOG),unigene is divided into 25 groups,with a total of 34323 unigene annotated.Among them,4641 genes(13.5%)are classified into translation process,ribosome structure and biogenesis.Based on the Kyoto Encyclopedia of Genes and Genomes(KEGG)metabolic pathway the genes are involved,the genes are divided into five branches:cell process,environmental information processing,genetic information processing,metabolism and organic system.Among them,the genes related to metabolic process accounted for the largest proportion,into which 23188 genes were classified,accounting for 61.1%of the total.In the process of metabolism,there are 3483 sugars metabolism genes,accounting for 15%,including starch and sucrose metabolism,gluconeogenesis.In addition,there are relatively more unigenes in ribosome and ribosome biosynthesis.According to Padj<0.05 and|log₂foldchange|>1,a total of 4959 differentially expressed genes were screened,including 2144 up-regulated genes and 2815down-regulated genes.Through the analysis of different genes by different ways,cell components and metabolic processes are the main processes,including the biosynthesis and catalytic activity of amino acids.Salt treatment had a great effect on the photosynthesis of S.glauca,in which the synthesis of related proteins in photosystem I and II decreased,and the synthesis of chlorophyll and light harvesting proteins decreased.By comparing unigene of S.glauca with the whole protein sequence of Arabidopsis thaliana though local blast.The results showed that some key genes in salt tolerance mechanism of S.glauca were found,including profilin and AQPs,which were up-regulated.In the process of ion localization,both the Sodium:proton antiporter and H^[+]-ATPase genes were up-regulated.As to osmoregulation,in addition to the accumulation of proline and betaine,the up-regulation of KAT gene and high affinity K⁺transporter gene,and the enhancement of the ability to absorb K⁺and other inorganic ions also play an important role.In the antioxidant system,catalase and glutathione transferase scavenging oxygen free radicals,GST gene up-regulation promotes the synthesis and accumulation of related enzymes in the antioxidant system.All of the above plays an important role in S.glauca dealing with salt stress.Finally,the actin gene(Cluster-7801.53944)was selected as the internal reference gene by comparing the unigene of S.glauca with the common internal reference gene of A.thaliana by local blast.The genes to be verified were detected by480 fluorescent real-time quantitative PCR,and the results of the analysis were consistent with the sequencing results.