The Selection And Functionalanalysis Of Abiotic Stress Relevant WRKY Transcription Factors From Hexaploid Wheat

As an important gene family,WRKY transcription factors could response to abiotic stress through regulating the expression of relevant genes at the transcriptional level.In this study,the genome databases of Triticum aestivum was used as the data resource.Local blast was performed between the WRKY transcription factors from Triticum urartu,Aegilops,and the sequences from Triticum aestivum.Through the identification in PFAM and SMART,a total of 123 TaWRKY transcription factors with full-length CDS was selected.And one WRKY transcription factor,TaWRKY146,was found to be up-regulated under abiotic stresses.The drought tolerance of TaWRKY146 over-expression lines was significantly improved All the detailed results were listed as following conclusions.1.The local blast was performed between the WRKY transcription factors from Triticum urartu,Aegilops,and the sequences from Triticum aestivum.818 similar sequences were got and submitted to PFAM and SMART platform.Finally,a total of 123 WRKY transcription factors with full-length CDS were got.These WRKY transcription factors were further made blast analyze with the abiotic related WRKY transcription factors in Arabidopsis thaliana and rice,and 18 abiotic related TaWRKYs was obtained for further research.2.The amino acid sequence of WRKY conserved domains of Triticum aestivum and Arabidopsis were made multiple alignment by Clustal X.The neighbor-joining method were used to construct unrooted phylogenetic tree with MEGA 5.10 software.All TaWRKYs were divided into three major groups:GroupⅠ,Ⅱ and Ⅲ,with 26,72 and 25 TaWRKYs respectively.TaWRKYs were distributed in different chromosomes broadly and unevenly.There are much repetitive TaWRKYs between A,D,B genomes.3.The gene expression pattern of 18 abiotic stress relevant TaWRKYs TFs performed by qRT-PCR and found that the gene expression pattern was different in leaf and root.The expression of Traes₁AL₀404BC790,Traes₃AL₁40B829CB,Traes₇DL_A9EF00572,Traes₁DL₅BAB0B6BC,Traes₁AL_F64E07A92 was up-regulated under abiotic stress,which showed signally relevant with abiotic stress.4.Traes₇DL_A9EF00572,which was named TaWRKY146,was up-regulated under ABA and H2O2 treatment,which showed that TaWRKY146 works rely on ABA signal transduction pathway.5.TaWRKY146 was cloned and sequenced.It belongs to Group Ⅲ and have a WRKY domain and a Zinc-finger motif of C2 HC.The result of qRT-PCR showed that TaWRKY146 plays an important role in wheat responding to abiotic stress.6.The over-expression lines of TaWRKY146 was obtained to launch stress expermient.Under the drought,the MDA content of over-expression lines was lower than wild line,but the proline and soluble sugar content was higher than wild line.These data showed that the over-expression of TaWRKY146 could reduce the damage of drought through the regulation of protective enzyme,which is important to stabilize internal environment of cell.7.The stomatal conductance,transpiration rate,leaf temperature was measured after drought stress to over-expression lines for three weeks.The result indicted that over-expression lines can deduce stomatal conductance and transpiration rate to maintain moisture and resist drought.