Screening Of Salt-Tolerant Mutants In Maize And Molecular Characterization Of ZmWRKY74

Maize is one of the most important crops in the world.Soil salinity is the main factor that can adversely affect maize growth,development and productivity.It is therefore imperative to study the molecular mechanism of salt tolerance in maize,to identify new tolerant genes,and to breed new cultivars with tolerance to salt stress.Stuides showed that transcription factors have important functions on improving crop tolerance to various stresses.WRKY transcription factor has become a hot spot in the gene function research.Members of this gene family are known to have diverse functions in regulating plant growth,development and response to biotic and abiotic stresses.Based on the maize EMS mutant library,this study screened 9 wrky mutants for their salt tolerance,and studied the molecular characteristics of candidate salt-stress-related gene ZmWRKY74.The results were as follows:1.Using maize B73 inbred line as the material,the appropriate Na Cl concentration for salt tolerance screening of maize seedling was determined to be 300 mmol/L.2.Based on the above screening concentration,salt tolerance screening was conducted for 9 copies of wrky mutants obtained by EMS mutagenesis,and finally 4 copies of salt tolerance mutants were obtained,namely E381,E481,E1180 and T136.Three moderately salt-tolerant mutants were E1065,E304 and E1728,respectively.2 salt sensitive mutants E307 and E621.3.Based on the salt screening results of maize mutants,ZmWRKY74 was selected as a candidate gene for further molecular characterization.First,the gene was cloned from maize B73 inbred line.The sequence analysis showed that the CDS length of ZmWRKY74 gene was 1227 bp,encoding 408 amino acids,and the relative molecular mass(Mw)of its coding protein was predicted to be 42.835 k Da,and the theoretical isoelectric point(p I)was 6.50.4.The analysis of tissue expression patterns showed that ZmWRKY74 gene had different expression levels in the selected 7 representative tissues.The expression levels of ZmWRKY74 gene were relatively high in roots,leaves,bracts and fruit ears,among which the expression levels in bracts were the highest.However,the expression level in stem,filament and tassel was relatively low,and the expression level in embryo and endosperm was very low,and almost no expression.5.Yeast hybridization experiments showed that ZmWRKY74 protein had transcriptional self-activation activity in yeast.6.The results of subcellular localization showed that ZmWRKY74 protein was located in the nucleus.