Genetic engineering of Mycobacterium leprae to Glow and Grow

项目摘要/摘要 在人类历史上，麻风病一直困扰着人类，至今仍困扰着200万至300万人。 发展中国家。尽管麻风杆菌是第一个被报道的细菌 与人类疾病相关的是，它从未在人工培养基上培养过。它可以是 生长在裸鼠的脚垫上，或系统地生长在九条带状的螳螂中。带着一代人 在14天的时间内，测试药物方案通过计算抗酸杆菌来执行，该抗酸杆菌是 既费力又费时。麻风杆菌DNA序列与麻风杆菌DNA序列的比较。 结核病表明，无法在人工培养基上培养麻风杆菌可能是由于 不能合成蛋氨酸、泛酸和分枝杆菌蛋白。使用基因工具，我们有 为结核分枝杆菌开发的，我们假设麻风杆菌可以用噬菌体稳定转化- 以整合为基础的熟练质粒。我们计划将生物发光共振技术 能量转移(BRET)-纳米和近红外蛋白质进入质粒检测 变形金刚。此外，我们假设麻风杆菌与失踪者的转变 蛋氨酸和泛酸的生物合成基因和mycobactin转运蛋白将使M. 麻风病在人工培养基上生长。一种发光的麻风杆菌菌株将增强药物 发展和评估控制麻风杆菌的免疫机制的能力 感染。麻风杆菌的基因工程在人工培养基上生长将增强我们的 对导致专性细胞内寄生的进化过程的理解。

Project Summary/ Abstract Leprosy has plagued mankind throughout human history and still afflicts 2 to 3 million people in Developing Countries. Although the leprosy bacillus was the first bacterium reported to be associated with human disease, it has yet never been cultivated on artificial media. It can be grown in the footpads of nude mice or systemically in nine-banded armadillos. With a generation time of 14 days, testing drug regimens are performed by counting acid-fast bacilli which is laborious and time-consuming. The comparison of the M. leprae DNA sequence to M. tuberculosis suggests the inability to culture M. leprae on artificial media might result from the inability to synthesize methionine, pantothenate and mycobactin. Using genetic tools, we have developed for M. tuberculosis, we hypothesize M. leprae can be stably transformed with phage- based integration proficient plasmids. We plan to incorporate a Bioluminescent Resonance Energy Transfer (BRET)-Nanoluc and Near infra-red proteins into the plasmid to detect transformants. Furthermore, we hypothesize that the transformation of M. leprae with the missing methionine and pantothenate biosynthetic genes and the mycobactin transporters will enable M. leprae to grow on artificial media. An M. leprae strain that glows would enhance drug development and the ability to assess the immunological mechanisms that control M. leprae infections. The genetic engineering of M. leprae to grow on artificial media would enhance our understanding of evolutionary processes that lead to obligate intracellular parasitism.