Study On The Function Of Wheat NF-Y Transcription Factor Genes TaNFYB-A13 And TaNF-YC1 To Resist Plant Osmotic Stress

Osmotic stresses such as drought and salinity seriously affect the growth and development of plants,and improving the ability of crops to resist osmotic stress is of great significance for achieving the goal of high yield and quality.In this study,based on the wheat NF-Y family transcription factor genes TaNFYB-A13 and TaNF-YC1 in response to osmotic stress,their functions of mediating plants against drought and salt stress were studied.The main results are as follows.1.The molecular analysis of the target gene shows that the TaNFYB-A13 gene cDNA is 519 bp in length,519 bp in open reading frame,encoding 172 amino acid residues,including 27 acidic amino acids,19 basic amino acids,and a molecular weight of 18.958kD.The pI value is 6.97;TaNFYB-A13 contains a conserved HAP3 domain(43 aa-106 aa),which belongs to the NF-YB type transcription factor family;the results of phylogenetic analysis show that TaNFYB-A13 is associated with Arthropoda,rice,and short anthers The genes of the NF-YB family of wild rice,corn and other species have high consistency in sequence composition.The full-length cDNA of TaNF-YCl gene is 839 bp,of which 1～27 bp is 5’ non-coding region,795～839 bp is 3’ non-coding region,and the open reading frame is 768 bp,encoding 256 amino acid residues,of which acidic amino acid is 27,19 basic amino acids,molecular weight 28.442kD,pI value 5.12;TaNF-YC1 protein contains a conserved HAP5 domain(87 aa～183 aa),belonging to the NF-YC type transcription factor family bound by CCAAT box;TaNF-YC1 has high consistency with the NF-YC family genes of soybean,bermudagrass and corn and other species in terms of nucleotide sequence composition.2.Analysis of the expression characteristics of the tested genes under drought and salt stress.The results showed that the expression levels of TaNFYB-A13 and TaNF-YC1 in wheat roots all showed an upward trend.This indicates that the above genes may play important biological functions in mediating plant response to drought and salt stress.3.Adopt DNA recombination technology to establish the tobacco lines of genetically transformed test gene positive and antisense sequences,and study the function of transgenic lines to resist osmotic stress.The results showed that under salinity and drought treatment,compared with the wild type,the plants expressing TaNFYB-A13 and TaNF-YC1 showed significantly increased leaf area,increased chlorophyll content,increased dry and fresh weight,and plant growth Significantly enhanced;the rate of stomata closure accelerated under drought treatment.In contrast,tobacco plants with antisense expression of the tested genes underwent salinity and drought treatment,the plant growth became worse,chlorophyll content was reduced,and dry and fresh weights were reduced;the rate of stomata closure was slow under drought treatment.This indicated that the tested genes may respond to osmotic stresses such as drought by regulating stomatal closure.4.Semi-quantitative expression analysis of the protective enzyme gene of TaNFYB-A13 transformed tobacco strains under NaCl treatment showed that compared with the wild type,the expression levels of the sense lines NtCAT,NtCAT1;3,NtPOD1;2,NtSOD1,NtSOD2 Significantly increased,while the expression of antisense lines decreased.This indicates that TaNFYB-A13 may enhance the expression and activity of the protective enzyme gene by regulating the transcription of the gene encoding the cell protective enzyme,thereby enhancing the plant’s ability to resist osmotic stress.5.In order to determine the subcellular localization of TaNF-YC1,a TaNF-YC1-GFP fusion expression vector was constructed.Through expression in tobacco epidermal cells and fluorescence observation,it was found that TaNF-YC1-GFP protein fluorescence was located In the cell nucleus,it has the same regulatory function as its transcription factor.6.In order to identify whether TaNFYB-A13 and TaNF-YC1 proteins interact,using yeast two-hybrid technology,two decoy vectors TaNFYB-A13-PGBKT7 and TaNF-YC1-PGADT1 were constructed.After repeated screening of defective media SD-Leu-Trp and SD-Leu-His-Trp,it was proved that there is an interaction between TaNFYB-A13 and TaNF-YC1 protein.