【作者】 谢帆；

【导师】 何勇强；

【作者基本信息】 广西大学 ， 生物化学与分子生物学， 2004， 硕士

【摘要】 病原菌从寄主中获得营养是病原菌致病所必需的，碳素营养是所需营养的基本成分。在缺乏葡萄糖等六碳糖的环境中，微生物会通过糖异生途径利用非糖物质来合成葡萄糖以维持生长。我们实验室的实验结果已证明野油菜黄单胞菌中编码糖异生途径的两个关键酶的基因(ppsA基因和maeB基因)的突变体的致病力显著低于野生型菌株，这表明糖异生途径是野油菜黄单胞菌致病所必需的。六磷酸葡萄糖异构酶(PGI)是糖异生途径中另一个关键酶，在野油菜黄单胞菌中编码该酶的基因是pgi基因(Xcc 8004基因组中基因编号为XC2379)。为了明确该基因是否在糖异生途径中起重要作用及其与Xcc致病性的关系，我们用质粒定点整合突变方法构建了pgi基因的突变体。碳源利用检测结果表明，pgi突变体对多种碳源，例如柠檬酸、琥珀酸、延胡索酸乙酸、乳糖等的利用能力很差，这说明该基因在糖异生途径中起重要作用。我们用剪叶接种法在Xcc的寄主植物满身红萝卜上检测了pgi突变体的致病力，接种十天后，pgi突变体的病斑长度是3.6mm，而野生型的病斑程度是9.6 mm。这一结果证明pgi基因是黄单胞菌致病所必需的。以上结果结合本实验室前期已证实的ppsA和maeB基因是黄单胞菌致病所必需的结论，进一步说明了糖异生途径在黄单胞菌致病的重要性。更多还原

【Abstract】 The ability to acquire nutrients from the host is essential for a pathogen to cause disease. Among those nutrients, carbon nutrient is one of the most important elements. Like other organisms, pathogens may use gluconeogenesis to synthesize glucose from non-sugar compounds when there is not sufficient hexose in their niche. The importance of the gluconeogenesis in Xcc virulence have been demonstrated recently in our laboratory. Glucose-6-phosphate isomerase (PGI) is another key enzyme in gluconueogenesis. According to Xcc 8004 genome annotation, Glucose-6-phosphate isomerase was encoded by pgi gene (ORF XC2379 ). To determined the role of this gene in gluconeogenesis and in pathogenesis of Xcc, mutants of pgi gene were constructed by homology plasmid integration method. The carbon utilization ability of the resulting mutant was tested, results showed that the pgi mutant grow poorly in non-carbon MMX medium with acetate, citrate, lactose, pyruvate, succinate, fumarate, or malate as sole carbon source. This indicated that pgi gene is involved in gluconueogenesis. The virulence of pgimutant was tested on Chinses radish (Raphanus sativus L. var. radical us Pers.) by leaf clipping, ten days post-inoculation, lesion length with a mean of 9.9 mm was observed on leaves inoculated with the wild type strain, while mean lesion lengths of 3.6 mm were observed with pgi mutant. This result demonstrated that pgi gene is required for full virulence of Xcc. These results combined with the previously findings in our laboratory that ppsA and maeB, genes encoding key enzymes in gluconeogenesis, are required for Xcc virulence, strongly indicated that the gluconeogenic pathway is important for Xcc virulence.更多还原