Cloning and nucleotide sequence of the drug resistant genes and its application to rapid identification.

研究了结核分枝杆菌临床分离株对利福平和链霉素耐药的分子机制。我们利用PCR技术对RNA聚合酶β亚基（rpoB）、核糖体S12蛋白（rpsL）和16SrRNA （rrs）的编码基因进行了测序。得到以下结论：1、对64株日本临床分离株rpoB基因进行了测序。90%以上的RFP耐药菌株可通过遗传方法检测到。2、用PCR扩增了173株日本临床分离株的rpsL基因，并采用Mbo - II酶切法将其分为两组。60.0%的sm耐药菌株（200mug/ml）不携带Mbo II位点，而sm耐药菌株（20mug/ml）和sm敏感菌株携带Mbo II位点。通过测序，在aa88上发现了另一个突变。对47株日本临床分离株的rrs基因进行PCR扩增和测序。35.1%的sm耐药菌株携带点突变或帧移位插入。通过遗传方法可检出75%以上的SM抗性菌株。

The molecular mechanisms of resistance to rifampicin and streptomycin in Mycobacterium tuberculosis clinical isolates have been examined. We have sequenced genes encoding the beta subunit of RNA polymerase (rpoB), the ribosomal S12 protein (rpsL) and 16SrRNA (rrs) by using PCR technique. Following conclusions have been obtained.1, The rpoB genes of 64 Japanese clinical isolates have been sequenced. More than 90% of RFP resistant strain can be detected by genetical method.2, The rpsL gene of 173 Japanese clinical isolates have been amplified by PCR and classified by Mbo II restriction digestion into two groups. 60.0% of SM-resistant (200mug/ml) isolates did not carry Mbo II site, whereas SM-resistant (20mug/ml) isolates and SM-sensitive isolates carried it. Another mutation was found on the aa88 by sequencing. The rrs gene of 47 Japanese clinical isolates have been amplified by PCR and sequenced. 35.1% of SM-resistant isolates carried a point mutation or frame shift insertion. More than 75% of SM resistant strain can be detected by genetical method.