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目的分析铜绿假单胞菌野生型菌株PAO1及PA0861突变菌株基因表达谱,探讨铜绿假单胞菌中GGDEFEAL基因PA0861的生物学功能。方法以野生型铜绿假单胞菌PAO1及PA0861基因转座子插入失活的突变体菌株为材料,利用结晶紫染色方法进行细菌生物膜的表型分析;利用Microarray技术分析其基因表达谱的变化;采用半定量RT-PCR验证Microarray结果。结果 PA0861突变株生物膜的合成量在4、8、12h均高于PAO1(P<0.05),在12h时增加了近1倍;利用Microarray技术筛选出差异表达基因517个,其中319个上调,198个下调。生物分子功能注释和Pathway分类显示这些差异表达基因除涉及代谢过程中基本的物质变化和能量转换外,突变体PA0861中涉及铁载体生物合成基因的表达增高。半定量RT-PCR验结果一致。结论铜绿假单胞菌突株PA0861基因表达谱发生改变,其中铁载体生物合成基因高表达,提示c-di-GMP可能通过对铁载体的调控影响细菌生物膜的形成。
Objective To analyze the gene expression profiles of PAO1 and PA0861 mutant strains of Pseudomonas aeruginosa and explore the biological function of PA0861 gene of GGDEFEAL gene in Pseudomonas aeruginosa. Methods The mutant strains of wild-type Pseudomonas aeruginosa PAO1 and PA0861 were inserted into inactivated mutant strains, and the biofilm phenotypes were analyzed by crystal violet staining. The changes of gene expression profiles were analyzed by Microarray Microarray results were validated by semi-quantitative RT-PCR. Results The biosynthesis of PA0861 mutant was higher than that of PAO1 at 4,8 and 12 h (P <0.05), and almost doubled at 12 h. The number of differentially expressed genes was 517 (319 of them were up-regulated by Microarray) 198 down. Biomolecular functional annotations and Pathway classification revealed that these differentially expressed genes, in addition to the underlying material changes and energy transitions involved in the metabolism, increased the expression of the gene involved in iron-carrier biosynthesis in mutant PA0861. Semi-quantitative RT-PCR results are consistent. Conclusions The gene expression profile of PA0861 in Pseudomonas aeruginosa strain is changed. The high expression of biosynthesis gene in iron carrier suggests that c-di-GMP may affect the formation of bacterial biofilm through the regulation of iron carrier.