摘要
目的构建用于白念珠菌MXR 1基因敲除的载体质粒,并通过Ura-Blaster策略敲除MXR 1两条等位基因。方法分别扩增白念珠菌MXR 1基因ORF两侧上下游的片段,通过酶切与连接反应,将上下游片段分别插入到p5921质粒的hisG-URA 3-hisG盒两端,从而形成MXR 1敲除载体质粒pUC-MXR 1-URA 3。通过Ura-Blaster策略将载体质粒转染到白念珠菌RM 1000内,并采用PCR和Southern-blot杂交方法鉴定各步转染、复筛所得的阳性克隆。结果成功获得MXR 1基因缺失的菌株。结论 MXR 1基因缺失菌株的构建,有助于深入研究白念珠菌耐药机制。
Objective To construct the plasmid for MXR1 disruption, and knockout the MXR1 gene in Candida albicans.Methods Up and down streams of MXR1 gene ORF were amplified and inserted into the p5921 plasmid.MXR1 gene was disrupted according to the Ura-Blaster method. The constructed strains were identified by PCR and Southern-blot methods.Results The knockout plasmid pUC-MXR1-URA3 and MXR1 disruption strains were successfully constructed.Conclusions The MXR1 disruption strains are helpful for the study of the drug-resistance mechanism.
出处
《中国真菌学杂志》
CSCD
2013年第2期69-73,共5页
Chinese Journal of Mycology
基金
国家自然科学基金(30872276)
