摘要
以人型结核杆菌基因组 DNA 为模板,合成二段引物各20个碱基进行聚合酶链式反应(PCR)。经琼脂糖凝胶电泳证实,获得一条245bp 扩增带。PCR 检测的敏感性染色体基因组DNA 为1pg,菌悬液为13个活菌/ml。在特异性试验中,人型结核杆菌、牛型结核杆菌、BCG可见此扩增带。被试的其它14种抗酸菌以及变铅青链霉菌、大肠杆菌质粒 PUC19、星状诺卡氏菌、红球菌均未见该扩增带。54例肺结核痰标本3种方法检查的阳性率分别为:萋尼氏抗酸染色16.7%,培养法14.8%,PCR 37.0%。前2种检查方法分别与 PCR 比较,经统计学处理均有显著性差异(P<0.01)。12例非结核性肺部疾患痰标本抗酸染色和 PCR 均为阴性。结果表明,PCR 技术是快速、敏感、特异诊断结核病的方法。
The PCR was used to detect M.tuberculosis DNA sequences in uncultured clinical speci-mens.Two oligonucleotide primers with 20 bp each amplified target templat DNA of M tuber-culosis.Amplified DNA product was 245 bp which was identificated by agarose gel electrophore-sis.The sensitivity of detection of M tuberculosis genomic DNA and bacteria suspension byPCR was lpg and 13 viable bacteria cell/ml,respectively.In specificity exprements,only M.tu-berculosis,M.bovis and BCG were positive by PCR,but all other 14 Mycobacter(?)m tested,inclu-ding streptomyces lividansand E.coli plasmid PUC19 were negative.The sensitivity of detection of M.tuberculosis by PCR was determined by comparing thelast-acid staining and culture on total 54 sputum specimens of pulmonary tuberculosis and 12nontuberculosis lung disease.The positive rate of PCR in pulmonary tuberculosis were 37.0%,culture method showed only 14.8%,fast-acid staining were 16.7%.Nontuberculosis lung diseasewere negative.The results show that DNA amplification is a superior method with high degree of sensiti-vity and specificity for rapid diagnosis of pulmonary tuberculosis.
出处
《微生物学报》
CAS
CSCD
北大核心
1992年第5期364-369,共6页
Acta Microbiologica Sinica
基金
解放军总后勤部卫生部科研基金
