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目的研究结核杆菌复苏因子基因的复苏作用。方法PCR扩增结核杆菌复苏因子基因片段并克隆至表达载体pET30a中,IPTG诱导表达复苏因子蛋白,经树脂纯化后进行SDS-PAGE和Western blot分析;按不同浓度和不同组合进行休眠结核菌复苏培养。结果PCR扩增获得结核杆菌复苏因子基因片段,大小分别为1 200、900、500、500和450 bp,与理论值相符。重组质粒转化pET30a菌,IPTG诱导获得有活性的复苏因子蛋白,分子质量单位为36 ku。复苏实验证明,复苏因子蛋白有促结核杆菌复苏和生长作用,以低浓度组合效果最佳。结论结核杆菌复苏因子蛋白对休眠结核杆菌有促复苏作用。
Objective To study the recovery of Mycobacterium tuberculosis recovery factor gene. Methods Mycobacterium tuberculosis recovery factor gene fragment was amplified by PCR and cloned into the expression vector pET30a. The recombinant protein was induced by IPTG and purified by SDS-PAGE and Western blot analysis. The recovery and culture of resting Mycobacterium tuberculosis was performed at different concentrations and combinations . Results The Mycobacterium tuberculosis recovery factor gene fragment was amplified by PCR and the sizes were 1 200, 900, 500, 500 and 450 bp, respectively, which were consistent with the theoretical values. Recombinant plasmids were transformed into pET30a and induced by IPTG. The molecular weight unit was 36 ku. Resuscitation experiments show that the recovery factor protein has the role of Mycobacterium tuberculosis recovery and growth, with the best combination of low concentrations. Conclusion Mycobacterium tuberculosis recovery factor protein has the role of promoting the recovery of Mycobacterium tuberculosis.