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目的研究白假丝酵母菌液泡相关基因的功能,寻找白假丝酵母菌潜在的药物靶点。方法选取ORF19.2734、ORF19.6950、ORF19.6605、ORF19.2006和ORF19.5780等5个潜在液泡功能相关基因,通过同源重组的方法构建基因缺失菌。分别观察这些基因缺失菌酵母态及菌丝态生长、液泡形态以及对各种刺激的敏感性,在阳性表型的基础上进一步研究基因功能。结果 ORF19.6950、ORF19.6605、ORF19.2006和ORF19.5780基因缺失菌在酵母态及菌丝态生长、液泡形态等方面与野生菌SN152无明显差异。ORF19.2734基因缺失菌酵母态生长速率减慢,对酮康唑敏感性升高,但其液泡形态未见明显改变。RT-PCR显示,ORF19.2734基因缺失菌与野生菌SN152相比,MDR1、ERG11、ERG2基因表达量降低。结论 ORF19.2734基因通过升高MDR1、ERG11、ERG2基因表达,降低菌株对唑类药物的敏感性。ORF19.2734基因可能是抗白假丝酵母菌耐药性产生的潜在靶点。
Objective To investigate the function of vacuolar related genes of Candida albicans and to find potential drug targets of Candida albicans. Methods Five potential vacuolar function related genes, ORF19.2734, ORF19.6950, ORF19.6605, ORF19.2006 and ORF19.5780, were selected to construct gene deletion bacteria by homologous recombination. The yeast strains, mycelial growth, vacuolar morphology and sensitivity to various stimuli of these deleted yeast cells were observed respectively, and the gene function was further studied on the basis of the positive phenotype. Results The deletion of ORF19.6950, ORF19.6605, ORF19.2006 and ORF19.5780 in yeast and mycelial growth, vacuolar morphology and other aspects of the wild strain SN152 no significant difference. ORF19.2734 gene deletion yeast yeast growth slowed down, increased sensitivity to ketoconazole, but no significant changes in the vacuolar morphology. The results of RT-PCR showed that the expression of MDR1, ERG11 and ERG2 genes was lower in ORF19.2734 deletion than wild type SN152. Conclusion ORF19.2734 gene can reduce the sensitivity of the strain to azoles by increasing the expression of MDR1, ERG11 and ERG2 genes. The ORF19.2734 gene may be a potential target for resistance to Candida albicans.