【摘要】 目的应用膜反向斑点杂交技术快速检测结核分枝杆菌对乙胺丁醇(EMB)耐药性。方法设计与合成用于检测结核分枝杆菌耐EMB基因embB的寡核苷酸探针,点于硝酸纤维素膜上,与结核分枝杆菌临床分离株生物素标记的聚合酶链反应(PCR)产物进行反向斑点杂交,并与PCR-直接测序(PCR-DS)结果比较。结果27株结核分枝杆菌临床分离株中,14株敏感株膜反向斑点杂交结果与标准株完全相同;13株耐乙胺丁醇临床分离株检测到embB基因突变,均为306位密码子ATG→GTG、ATG→CTG、ATG→ATA、ATG→ATT和ATG→ATC突变,该突变与直接测序结果完全一致。结论膜反向斑点杂交技术可能成为检测部分结核分枝杆菌乙胺丁醇耐药基因型简便、快速的方法。更多还原

【Abstract】 [Objective] To study the rapid detection of resistance to ethambutol in mycobacterium tuberculosis by reverse dot blot hybridized technique.[Methods] We designed and synthesized the oligonucleotide probes of ethambutol-resistant genes(embB) and drop it on pyroxylin membrane.The target DNA fragments of M.tuberculosis clinical isolates were labeled with biotin by PCR amplification,and then hybridized with oligonucleotide probes on membrane.PCR-direct sequencing(PCR-DS) techniques were used as the control.[Result] The embB gene in 27 mycobacterium tuberculosis clinical isolates were analyzed.membrane hybridization in 14 sensitive strains were similar to H37Rv and 13 strains were positive hybridization.They were all mutations ATG→GTG,ATG→CTG,ATG→ATA,ATG→ATT和ATG→ATC at codon 306 of embB,which were consistent with those of DNA sequencing.[Conclusion] The membrane-reverse dot blot hybridized technique was simple and rapid and may be used to detect ethambutol-resistance of partial mycobacterium tuberculosis clinical strains.更多还原