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目的:用甲基磺酸乙酯(Ethylmethylsulfone,EMS)诱导D型沙眼衣原体突变,利用间接免疫荧光法筛选出突变菌株,为研究不同衣原体基因的功能提供实验依据。方法:将D型沙眼衣原体标准株接种Mc Coy细胞,加入EMS诱导突变,收集存活菌株,利用空斑实验进行衣原体的分离和纯化,并用不同衣原体蛋白单克隆抗体做间接免疫荧光实验筛选突变株。结果:用间接免疫荧光筛选经EMS作用的沙眼衣原体,筛选出三株包涵体形态偏小的菌株(56#、58#、95#),一株圆形包涵体的突变株(61#)和一株D413N表达阴性的突变菌株(83#)。结论:用EMS作为诱导剂诱导D型沙眼衣原体突变,并成功筛选出三种突变株。为寻找衣原体功能基因与衣原体表型之间的联系奠定了实验基础。
OBJECTIVE: To induce the mutation of Chlamydia trachomatis D with Ethylmethylsulfone (EMS) and to screen out the mutant strains by indirect immunofluorescence, which can provide experimental basis for studying the function of different Chlamydia trachomatis genes. Methods: The standard strain of Chlamydia trachomatis was inoculated with Mc Coy cells. EMS mutagenesis was added to induce the survivin strain. Chlamydial isolation and purification were performed by plaque assay. Mutants were screened by indirect immunofluorescence assay using different monoclonal antibodies against Chlamydia pneumoniae. Results: Indirect immunofluorescence was used to screen for C. trachomatis with EMS, and three strains (56 #, 58 # and 95 #) with smaller size and one circular inclusion body (61 #) and A D413N negative mutant strain (83 #). Conclusion: The mutation of Chlamydia trachomatis type D was induced by EMS as an inducer, and three mutant strains were successfully screened. It laid the experimental foundation for finding the relationship between Chlamydia functional genes and the phenotype of Chlamydia.