Study On Salt Tolerance Function Of RcCDPKs Gene In Ricinus Com Munis L.

Soil salinization hinders plant development and reduces crop yield,and is one of t he most important abiotic stresses for plant production.The use of genetic engineering means to breed salt-tolerant varieties of crops can effectively improve and utilize salin e soils.Salt-treated castor seeds were used as experimental materials,and the calcium-d ependent protein kinases（CDPKs）RcCDPK29 and RcCDPK33,which were significan tly up-regulated by the transcriptomic data of salt stress in castor,were successfully clo ned and characterized.The functions of RcCDPK29 and RcCDPK33 were analyzed by Agrobacterium tumefaciens-mediated transformation of Arabidopsis thaliana and cast or with different levels of salt stress treatment.This study laid the foundation for breed ing new salt-tolerant germplasm of castor,and the main results obtained are as follows:1.Successful cloning of RcCDPK29 and RcCDPK33 genes.The ORF of RcCDPK33 is 1602 bp and encodes 533 amino acids,including a Ser/Thr protein kinase catalyt ic domain at positions 205-217 and（390-418,426-454,462-490,497-525）.There are als o eight Ca²⁺binding sites（399,401,403,410,435,437,439,446）;the ORF of RcCDPK29 is 1590 bp,encodes 528 amino acids and contains a Ser/Thr protein kinase catalytic domain at positions 81-338 and four Ca²⁺-binding EF-hand-type structural domains（385-413,421-449,457-485,492-520）.2.The expression of RcCDPK33 and RcCDPK29 in different tissues of castor un der different levels of salt stress was analyzed by q RT-PCR,and the results showed tha t both RcCDPK33 and RcCDPK29 had the highest expression in stems,and the expres sion of RcCDPK33 peaked at 24h of salt treatment in stems,and RcCDPK29 peaked a t 12 h of treatment.peak expression.3.RcCDPK33 and RcCDPK29 overexpression vectors were constructed,and Ara bidopsis and castor were transformed by Agrobacterium rhizogenes-mediated transfor mation of RcCDPK33 and RcCDPK29.Phenotypic analysis of transgenic Arabidopsis revealed that the seed germination rate and seedling root length were highly significan t higher than those of wild type.The results of physiological indexes measured in trans genic Arabidopsis and castor under salt stress reveal that SOD and CAT intensities are significantly higher than those of wild type,and MDA content is highly and significan tly down-regulated,indicating that both castor and Arabidopsis overexpressing RcCD PK33 and RcCDPK29 exhibit strong salt tolerance.4.The RcCDPK29 gene with high up-regulated expression ploidy was selected fr om the two genes to construct a CRISPR/Cas9-edited vector,and the CRISPR/Cas9-R c CDPK29 castor 9 plants were obtained by transformation of RcCDPK29 mediated by Agrobacterium tumefaciens.The physiological indexes of CRISPR/Cas9-edited RcCD PK29 gene plants were measured,and the results showed that the SOD and CAT activi ties were extremely significantly lower and the MDA content was extremely significan tly higher than that of the wild type.