【摘要】：目的：克隆結核分枝桿菌rpoB、katG常見基因突變位點,為核酸薄膜導流雜交技術快速檢測臨床標本結核分枝桿菌rpoB、katG基因突變提供陽性對照,以助判斷臨床標本結核分枝桿菌有無耐藥。 方法：前期研究發(fā)現本地區(qū)結核分枝桿菌rpoB、katG基因常見的4個突變位點和6個突變形式,選擇6株含有上述突變特征的臨床分離株,PCR擴增其rpoB、katG基因片段進行T-A克隆,采用HindⅢ和EcorⅠ雙酶切、PCR擴增及重組質粒測序鑒定重組質粒。核酸薄膜導流雜交技術檢測此6個臨床分離株的rpoB、katG基因片段。 結果：構建了6個含有rpoB、katG基因不同突變類型的重組質粒,三種方法鑒定明確重組質粒含有正確序列。核酸薄膜導流雜交技術檢測6個臨床分離株的rppB、katG基因片段結果顯示在預期的位置顯示陽性斑點。 結論：重組質�？梢宰鳛楹怂岜∧Я麟s交技術檢測臨床標本時提示結核分枝桿菌有無耐藥的穩(wěn)定的陽性對照。
[Abstract]:Objective: to clone the common mutation sites of Mycobacterium tuberculosis rpoBhkatG gene, and to provide a positive control for rapid detection of rpoB katG gene mutation in clinical specimens by membrane diversion hybridization, in order to help determine the resistance of Mycobacterium tuberculosis in clinical specimens. Methods: four common mutation sites and six mutation forms of Mycobacterium tuberculosis rpoBnkatG gene were found in previous studies. Six clinical isolates with the above mutational characteristics were selected for T-A cloning by PCR amplification of the rpoBmkatG gene fragment of Mycobacterium tuberculosis. The recombinant plasmids were identified by Hind 鈪，

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