Stimulation of transposition of the Mycobacterium tuberculosis insertion sequence IS6110 by exposure to a microaerobic environment
Stimulation of transposition of the Mycobacterium tuberculosis insertion sequence IS6110 by exposure to a microaerobic environment
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DOI：
10.1046/j.1365-2958.1999.01539.x
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发表时间：
1999-09-01
影响因子：
3.6
通讯作者：
McFadden, J
中科院分区：
文献类型：
--
作者：
Ghanekar, K;McBride, A;McFadden, J
结核分枝杆菌特异性插入序列IS 6110/986由于在不同菌株间观察到的多重多态性而被广泛用作探针。为了研究IS 6110的转座作用，构建了一系列人工构建的含有IS 6110和卡那霉素抗性标记的复合转座子。将复合转座子插入到条件复制、温度敏感的大肠杆菌-分枝杆菌穿梭载体中，并导入M。在允许温度下，使转化体的菌苔在补充卡那霉素的琼脂上生长，随后通过转移到非允许温度来防止进一步生长。在正常的大气条件下，卡那霉素抗性乳头仅在孵育约5-6周后出现。然而，这些事件与转座子动员无关。相比之下，草坪暴露于48小时的微需氧休克产生卡那霉素抗性乳头后，只有6-14天。这些事件是通过将IS 6110复合转座子保守转座到M.染色体缺失，穿梭载体缺失。与其他IS 3家族元件一样，IS 6110的转座被认为是由翻译移码控制的。然而，我们无法检测到任何显着的移码内推定的移码网站的IS 6110和移码水平不受微需氧孵育。IS 6110的转座通过在降低的氧张力下孵育而被刺激的发现可能与IS 6110在M中的转座有关。结核病病灶内存在结核病。
The Mycobacterium tuberculosis-specific insertion sequence IS6110/986 has been widely used as a probe because of the multiple polymorphism observed among different strains. To investigate transposition of IS6110 a series of artificially constructed composite transposons containing IS6110 and a kanamycin resistance marker were constructed. The composite transposons were inserted into a conditionally replicating, thermosensitive, Escherichia coli-mycobacterial shuttle vector and introduced into M. smegmatis mc(2)155, Lawns of transformants were grown at the permissive temperature on kanamycin-supplemented agar and subsequently prevented from further growth by shifting to the nonpermissive temperature. Under normal atmospheric conditions, kanamycin-resistant papillae appeared after only about 5-6 weeks of incubation. However, these events were not associated with transposon mobilization. In contrast, lawns that were exposed to a 48 h microaerobic shock generated kanamycin-resistant papillae after only 6-14 days. These events were generated by conservative transposition of the IS6110 composite transposon into the M. smegmatis chromosome, with loss of the shuttle vector. In common with other IS3 family elements, transposition of IS6110 is thought to be controlled by translational frameshifting. However, we were unable to detect any significant frameshifting within the putative frameshifting site of IS6110 and the level of frameshifting was not affected by microaerobic incubation. The finding that transposition of IS6110 is stimulated by incubation at reduced oxygen tensions may be relevant to transposition of IS6110 in M. tuberculosis harboured within TB lesions.