| In this study,rice Tonghe 899(control strain P)and its stable high-generation salt-tolerant strain 2020-8014(strain A)and salt-sensitive strain 2020-8056(strain C)were selected by high energy heavy ion beam mutagenesis.Through the determination of the related phenotypes and physiological indexes of the three strains under normal conditions(M)and salt stress conditions(S),the physiological basis of salt tolerance was analyzed.Meanwhile,the expression levels of salt tolerance genes between the high generation strains and the control strains were analyzed by transcriptome sequencing,which provided scientific reference for the mining of salt tolerance genes and the breeding of salt-tolerant rice varieties.The main results are as follows:1.Phenotypic variation levels of plant height,fresh weight,root length and other agronomic traits between Tonghe 899 and two high-generation strains were significantly different under high salt treatment.Under salt stress,the fresh weight of Tonghe 899,strain A and strain C decreased by 31.13%,31.25% and 48.48% compared with normal condition.The plant height of Tonghe 899 was reduced by 27.24%,33.41% and 40.07%compared with normal condition.Compared with normal condition,the root length of Tonghe 899 decreased by 12.66%,strain A by 12.71% and strain C by 32.79%.2.By comparing the MDA content of Tonghe 899 and two high-generation strains under normal conditions and salt stress treatment,the MDA content of Tonghe 899 increased by 12.42%,9.66% in strain A and 17.32% in strain C.The increase of MDA content in strain A was the smallest,which suggested that strain A had the lowest degree of membrane system damage under salt stress,which might be one of the reasons for its salt tolerance.3.By measuring the contents of soluble protein,soluble sugar and proline of the three groups of materials under normal conditions and salt stress conditions,it was found that the contents of osmoregulatory substances in the body of the three strains would change when they were subjected to salt stress.The results showed that the soluble protein content of control strain P decreased by 6.86%,that of strain A by 1.14%and that of strain C by 13.69%.The soluble sugar content of strain P,strain A and strain C increased by 11.26%,20.70% and 10.62% respectively.The proline content of control strain P increased by 13.3 times,that of strain A by 16.2 times,and that of strain C by11.31 times.It was speculated that strain A could control the osmotic pressure inside and outside the cell by making better use of soluble sugar,soluble protein,proline and other osmoregulatory substances,thus reducing water loss,reducing the damage caused by salt stress,and expressing strong salt tolerance.4.By comparing the SOD and POD activities of the three groups of materials under normal conditions and salt stress,it was found that the SOD and POD activities decreased in different degrees under salt stress conditions,among which the SOD activity of control strain P decreased by 13.11%,that of strain A decreased by 4.41%,and that of strain C decreased by 7.71%.The POD activity of control strain P was decreased by 26.96%,that of strain A by 11.91% and that of strain C by 31.62%.The results showed that strain A could express salt tolerance by better maintaining SOD and POD activities under salt stress.5.GO enrichment analysis was conducted on the differentially expressed genes obtained from the three groups of materials under normal conditions and salt stress conditions.The results showed that,under salt stress conditions,the differentially expressed genes of strain A and control strain P were mainly enriched in two aspects of molecular function and biological process.In biological processes,Differentially expressed genes are enriched in ubiquitine protein catabolism,modified-dependent protein catabolism,modified-dependent macromolecular metabolism,cellular macromolecular catabolism,cellular protein catabolism,hydrolyzed proteins involved in cellular protein catabolism,organic nitrogen compound catabolism,protein catabolism,etc In various metabolic processes.Under salt stress,the differentially expressed genes of strain C and control strain P were mainly in two aspects: molecular function and biological process.Differentially expressed genes were significantly enriched in the hydrolase activities of glycosylated bonds,hydrolase activities of O-glycosylated compounds,peptidase inhibitors,peptidase regulatory activities,endopeptidase regulatory activities,endopeptidase inhibitor activities and other processes.In the biological process,the reaction to acidic compounds,the reaction to water,carbohydrate metabolism,the reaction to inorganic substances and other processes are significantly enriched.KEGG pathway analysis showed that the differentially expressed genes of strain A and control strain P were mainly concentrated in the organic system,metabolism and protein hierarchy pathways under salt stress,and the organic system process was the main pathway.Organic system processes include organic system,environmental adaptation,plant-pathogen interaction and other pathways.The metabolic processes include glutathione metabolism and other amino acid metabolism pathways.The protein hierarchy process includes cytochrome P450 and transporter pathways.The differentially expressed genes of strain C and control strain P were mainly concentrated in the three main pathways of environmental information processing,metabolism and protein hierarchy,and the metabolic process was the main pathway.The metabolic process includes metabolism,other amino acid metabolism,carbohydrate metabolism,glutathione metabolism,galactose metabolism,tyranine metabolism,amino acid metabolism and other pathways.The process of protein hierarchy includes transporter and cytochrome P450 pathways.Environmental information processing includes plant-Mark signal path and other pathways.6.The transcription factors of the three groups of materials were analyzed under normal conditions and salt stress conditions,and the gene expressions of the larger families of transcription factors were obtained.The results showed that the majority of transcription factors in the six groups of materials were located in the AP2,WRKY,b HLH,b ZIP and MYB families.The results showed that the expression of different transcription factor-related genes changed to different degrees under salt stress,suggesting that they play a role in rice resistance to stress.7.Hormone regulatory genes in Ethylene were analyzed in ethylene under normal and salt stress conditions.The results showed that abscisic acid(ABA),growth hormone(IAA),ethylene(Ethylene)related genes were all changed in ethylene.According to the analysis results of hormone-related genes in the test materials,the expression differences of the three hormone-related genes in the three groups of materials were small under normal conditions and salt stress conditions,while the gene expression differences were large under normal conditions and salt stress conditions,suggesting that the expression of the three hormone-related genes may play a role in the process of salt tolerance in rice.8.The ion channel regulatory genes of the three groups of materials under normal conditions and salt stress conditions will be analyzed.The results showed that there was little difference in the expression of ion channel regulatory genes between the three groups of materials under normal conditions and salt stress conditions,while there was a big difference in the expression of gene under normal conditions and salt stress conditions,suggesting that ion channel regulatory expression may play a role in the process of salt tolerance in rice. |
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