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【目的】确定玉米大斑病菌转录因子StSte12在基因组中的位置,分析目的蛋白质StSte12的结构特征;明确StSTE12在病菌不同发育时期的表达情况;筛选可能受StSte12转录因子调控的基因,分析其匹配序列的结构特征,并比较它们在病菌的中表达情况。【方法】利用生物信息学方法,确定StSte12在玉米大斑病菌基因组中的位置并解析目的蛋白质StSte12的结构特征;利用Western blot技术分析StSTE12在病菌不同发育时期的表达规律;根据酿酒酵母Ste12转录因子的下游调控基因,筛选病菌中可能受StSte12调控的功能基因;并利用生物信息学方法得出下游调控基因的结合位点;利用RT-PCR技术,分析这些基因在StSTE12 RNAi转化子与野生型菌株中的表达情况。【结果】StSTE12的ID为105656,该基因位于scaffold_13正链的1061747-1064127位置;该蛋白具有Ste12-like蛋白特有的保守结构域(STE homeodomain和ZnF_C2H2锌指结构)及空间结构;StSTE12在附着胞时期表达量最大;筛选了部分StSte12调控基因,与StSte12的匹配序列为TC GAAAC AG,中间部分GAAAC非常保守;StKAR5在野生型中表达量小于StSTE12 RNAi转化子,StBEM2、StBUD8、StCHS7相反;StRAX2的表达量在野生型和转化子中一致。【结论】StSte12具有Ste12-like所特有的DNA结合结构域和空间结构;该蛋白在附着胞发育中发挥重要功能;筛选了部分StSte12的调控基因,其匹配序列可概括为TC GAAAC AG,且证实了这些基因受StSte12调控。
【Objective】 The objective of this study was to determine the position of StSte12, a transcription factor of S. turcica, in the genome and to analyze the structural characteristics of the target protein StSte12. The expression of StSTE12 in different developmental stages of the bacteria was determined. The genes that may be regulated by StSte12 transcription factor were screened and their matching sequences The structural characteristics of the bacteria were compared and their expression in the bacteria was compared. 【Method】 The bioinformatics method was used to determine the position of StSte12 in the genome of S. turcica and to analyze the structural characteristics of the target protein StSte12. Western Blot was used to analyze the expression pattern of StSTE12 in different developmental stages of S. cerevisiae. Of the downstream regulatory genes, screening of pathogens may be regulated by StSte12 function genes; and use of bioinformatics methods to obtain the downstream regulatory genes binding sites; using RT-PCR analysis of these genes in StSTE12 RNAi transformants and wild-type strains In the expression of the situation. 【Result】 The ID of StSTE12 was 105656, located at the position 1061747-1064127 of the positive strand of scaffold_13, which possessed the unique conserved domains of STE homeodomain and ZnF_C2H2 and the spatial structure of St12-like protein. The expression level of StKAR5 in wild type was less than that of StSTE12 RNAi transformants, StBEM2, StBUD8 , Contrary to StCHS7; StRAX2 expression was consistent between wild type and transformants. 【Conclusion】 StSte12 possesses Ste12-like specific DNA binding domain and spatial structure. The protein plays an important role in appressorium development. The partial StSte12 regulatory genes are screened and their matching sequences can be summarized as T C GAAAC A G, and confirmed that these genes are regulated by StSte12.