DNA指纹技术方法学的建立及在结核分支杆菌菌株鉴定中的应用

Methodology of DNA fingerprinting and its application in identification of M. tuberculosis

目的探讨结核分支杆菌ＤＮＡ指纹技术的方法学及其在结核分支杆菌菌株鉴定中的应用。方法以结核分支杆菌染色体ＤＮＡ插入序列ＩＳ６１１０为基础，根据不同来源的结核分支杆菌菌株ＤＮＡ相异的ＩＳ６１１０拷贝数和相对分子质量，应用增强化学发光标记技术对结核分支杆菌ＤＮＡ酶切产物进行杂交和检测。结果不同来源的５０例结核患者临床分离株具有相异的ＤＮＡ指纹图谱。同一患者痰和膝关节脓培养阳性的结核分支杆菌菌株ＤＮＡ指纹图谱有较大的差异。人工诱导的结核分支杆菌Ｈ３７Ｒｖ氧氟沙星耐药株和敏感株有一致的ＤＮＡ指纹图谱。

Objective To explore methodology of DNA fingerprinting technique and its application in identification of strains of M. tuberculosis. Methods This research was based on different copy numbers and location of chromosome DNA IS6110 in M. tuberculosis genome. After DNA of M. tuberculosis were cut by endonuclease PvuII, the products were transferred to nylon membrane, then detected and hybridized by enhanced chemiluminescent(ECL)labeling techniques. Results Fifty clinical isolates from different TB patients had unidentical DNA fingerprinting patterns. DNA fingerprinting patterns showed a great difference from sputum and pus specimens of the same patient. H 37 R v ofloxacin resistant and sensitive strains had an identical DNA fingerprinting pattern. Conclusions Identification of M. tuberculosis strains is feasible by DNA fingerprinting technique.