| Sorghum,as an important cereal crop,is widely used and cultivated all over the world.Sorghum will face the stress of adverse factors such as drought and high temperature during the growth process,resulting in undesirable yield and quality.Due to the low efficiency and long breeding cycle of traditional breeding methods,it is great significance to explore how to improve the resistance of sorghum using modern biotechnology and molecular biology methods.As an important transcription factor gene,WRKY has important biological functions in regulating plant growth and development,responding to biological and abiotic stresses.Thus,this study applied bioinformatics methods to identify 97 WRKY transcription factors in the whole sorghum genome,and conducted detailed classification,conserved motif and phylogenetic relationship analysis.At the same time,used transcriptome sequencing to analyze the expression of related genes which under drought stress,and these genes were subjected to Go functional annotation and KEGG metabolic pathway analysis.Sb WRKY transcription factors responding to drought stress were screened and their functions were analyzed,the Sb WRKY28 gene was cloned.The main results are as follows:1.The 97 Sb WRKY transcription factors were divided into three categories,of which 11 were in group Ⅰ,70 in group Ⅱ,which could are subdivided into five subgroups a to e,and 16 in group Ⅲ.The WRKY heptapeptide domain of 16 Sb WRKY transcription factors were mutated or the zinc finger structure was special.In addition,the phylogenetic relationship shows that group Ⅱ and Ⅲ are derived from group Ⅰthrough long-term variation and evolution.2.Through transcriptome sequencing analysis,a large number of differentially expressed genes were screened which responding to drought stress.These genes were mainly involved in three metabolic pathways of carbon metabolism,energy metabolism and photosynthesis.They coordinated the three major metabolic pathways to deal with drought stress by enhancing the utilization rate of carbon and energy and regulating photosynthetic intensity in sorghum.3.Obtained 4 WRKY transcription factors which response to drought stress.Through Go metabolic pathway analysis found that they can respond to drought stimuli and enhance stress resistance ability by activating the expression of drought-related genes and enhancing the transcriptional regulation of drought-related genes.The expression patterns analysis of 4 genes found that Sb WRKY24 was rapidly down-regulated under drought stress,while the other three were up-regulated genes,especially the expression level of Sb WRKY28 induced by drought was higher than Sb WRKY35 and Sb WRKY79,which indicate that Sb WRKY28 was more sensitive to drought stress.It is speculate that Sb WRKY28 may be involved in the regulation of drought resistance in sorghum.4.Cloned into the Sb WRKY28 gene,which has a CDS of 651 bp and encodes 216 amino acids.The p BI121-Sb WRKY28 plant over expression vector was constructed,and the expression vector was transformed into Agrobacterium to prepare for further verification of Sb WRKY28 gene function.5.The molecular weight and isoelectric point of Sb WRKY28 protein are 23.76 kd and 5.57,respectively.The Sb WRKY28 protein belongings to the WRKY family Group Ⅱb.Bioinformatics analysis revealed that the Sb WRKY28 protein is a hydrophilic,non-secreted protein with no signal peptide structure,no transmembrane domain,with multiple phosphorylation sites,it can play signal transduction and perform function in cells. |
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