Mokrousov Igor, Narvskaya Olga, Limeschenko Elena, Otten Tatiana, Vyshnevskiy Boris
Laboratory of Molecular Microbiology, Pasteur Institute, 14, Mira Street, St. Petersburg 197101, Russia.
J Clin Microbiol. 2002 May;40(5):1617-20. doi: 10.1128/JCM.40.5.1617-1620.2002.
We describe a multiplex allele-specific (MAS)-PCR assay to detect simultaneously mutations in the first and third bases of the embB gene codon 306ATG. These mutations are known to confer ethambutol (EMB) resistance in the majority of clinical Mycobacterium tuberculosis isolates worldwide. The mutated bases are revealed depending on the presence or absence of the respective indicative fragments amplified from the embB306 wild-type allele. Initially optimized on purified DNA samples, the assay was tested on crude cell lysates and auramine-stained sputum slide DNA preparations with the same reproducibility and interpretability of the generated profiles in agarose gel electrophoresis. Since EMB resistance is generally linked to multiple-drug resistance (MDR), the MAS-PCR assay for EMB resistance detection can be used in clinical laboratory practice in areas with a high prevalence and a high transmission rate of MDR-EMB-resistant tuberculosis.
我们描述了一种多重等位基因特异性（MAS）-PCR检测方法，用于同时检测embB基因密码子306ATG第一位和第三位碱基的突变。已知这些突变在全球大多数临床结核分枝杆菌分离株中赋予乙胺丁醇（EMB）抗性。根据从embB306野生型等位基因扩增出的相应指示性片段的有无来揭示突变碱基。该检测方法最初在纯化的DNA样本上进行了优化，随后在粗细胞裂解物和金胺染色的痰涂片DNA制剂上进行了测试，在琼脂糖凝胶电泳中生成的图谱具有相同的重现性和可解释性。由于EMB抗性通常与多重耐药性（MDR）相关，用于检测EMB抗性的MAS-PCR检测方法可用于耐多药-耐EMB结核病高流行率和高传播率地区的临床实验室实践。
