Cloning And Functional Analysis Of Aldehyde Ketone Reductase Gene (MSAKR1) In Response To Salt Stress In Alfalfa

The plant aldo-keto reductases(AKR)superfamily plays an important role in adversity stress.In order to analyze and verify its role in Medicago,the AKR gene of alfalfa was obtained by homologous cloning method,and its function was verified by fluorescence quantitative analysis and transgenic method,which provides theoretical basis for improving salt tolerance of legumes.The main results are as follows:1.Thirty members of AKR family of Medicago truncatula were obtained.Their gene properties,protein basic properties,conserved motifs and cis acting elements were analyzed.The results showed that the expression levels of MtAKR 1,MtAKR5,MtAKR 11,MtAKR 14,MtAKR 20 and MtAKR 29 in family members treated with PEG,NaCl and ABA were significantly increased,which could be used as candidate genes for the stress regulation of aldo-keto reductases family.2.The homologous gene of MtAKRl in Medicago sativa was selected,and its cds sequence,amino acid composition,protein secondary structure and tertiary structure were analyzed.The expression vector pCAMBIA3301-MsAKR was constructed and transformed into Arabidopsis thaliana and alfalfa(ZhongMu NO.1)respectively.The expression of MsAKR1 gene in leaves was 30 times higher than that in roots.The results of subcellular localization in Arabidopsis thaliana protoplast showed that MsAKR1 was expressed in chloroplast.3.RT-qPCR analysis of Alfalfa treated with low temperature(4?),NaCl(200 mM)and PEG 6000(50 g·L-1)showed that the expression of MsAKR1 was up-regulated in roots and leaves.MsAKR may play a role in resistance to these stresses.4.18 transgenic Arabidopsis thaliana plants with MsAKR1 were obtained at the genomic level,and two transgenic lines were selected to study the stress resistance of Arabidopsis thaliana.The results of vertical plate showed that the growth potential of transgenic Arabidopsis thaliana was better than WT,and 150 mM and 200 mM mannitol concentration was the better select concentration.The results of treatment of wild type,mutant and T3 transgenic Arabidopsis thaliana with 200 mM NaCl solution showed that the growth potential and plant height of transgenic lines were better than those of WT and mutant materials.The results showed that the content of CAT was higher than that of WT and mutant materials,and the content of MDA was lower than that of WT and mutant materials.5.30 overexpression MsAKR lines of alfalfa were obtained at the genomic level,.Two lines were selected to study the stress resistance of alfalfa.The results showed that the growth and leaf color of transgenic plants were better than those of WT.The activity of CAT was higher than that of WT,and the content of MDA was lower than that of WT.The results showed that MsAKR1 may play an important role in the regulation of salt tolerance and other adversity in alfalfa.