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目的用实时荧光定量PCR确定伯氏疏螺旋体感染小鼠模型不同组织的病原体载量。方法培养低传代伯氏疏螺旋体至对数生长期,稀释为1×105/ml。为建立伯氏疏螺旋体感染小鼠模型,于每只小鼠皮内注射菌液100μl,在证实感染成功后,于第12天和第18天分别取不同组织,提取总DNA,用实时荧光定量PCR分别测定组织中的螺旋体flaB基因拷贝数,并标准化为每106β-肌动蛋白所对应的flaB拷贝数(螺旋体数)。对不同组织的螺旋体载量进行统计学处理,确定不同组织螺旋体载量差异是否有统计学意义。结果在所检测的4种代表性组织中,膀胱螺旋体载量在两个典型时间点均最高,皮肤和关节次之,心脏最低。结论伯氏疏螺旋体感染小鼠后,不同组织螺旋体载量差异有统计学意义。组织螺旋体载量与组织损伤程度无密切关系。
Objective To determine the pathogen load in different tissues of mice infected with Borrelia burgdorferi by real-time fluorescence quantitative PCR. Methods Low-passage Borrelia burgdorferi was cultured to logarithmic growth phase and diluted to 1 × 10 5 / ml. In order to establish a mouse model of Borrelia burgdorferi infection, each mouse was injected intradermally with 100 μl of bacterial liquid, and after confirming the successful infection, different tissues were taken on the 12th day and the 18th day respectively to extract total DNA, and real-time fluorescence quantitative PCR was used to determine the number of flaB gene copies in the tissue and normalized to the number of flaB copies (spirochetes) per 106β-actin. Spiral load of different tissues were statistically analyzed to determine whether there was significant difference in the load of spirochetes among different tissues. Results Among the four representative tissues tested, the load of cystospiral was the highest at two typical time points, followed by the skin and joints, and the lowest in the heart. Conclusion After the mice were infected with Borrelia burgdorferi, there were significant differences in the volume of spirochetes in different tissues. Tissue spiral volume and tissue damage is not closely related.