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Genetic engineering of Mycobacterium leprae to Glow and Grow

麻风分枝杆菌发光和生长的基因工程

基本信息

批准号：
10526890
负责人：
WILLIAM Robert JACOBS
金额：
$ 25.2万
依托单位：
ALBERT EINSTEIN COLLEGE OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-05-20 至 2024-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10526890
关键词：
ATP-Binding Cassette Transporters Achievement Acid Fast Bacillae Staining Method Anabolism Animal Model Bacillus Bacteria Bacteriophages Bioluminescence Carbon Cell Wall Cells Communities Complement Consumption Cosmids DNA DNA Sequence Data Detection Developing Countries Development Diagnosis Disease Electroporation Energy Transfer Engineering Enzymes Evolution Family Dasypodidae Fatty Acids Fireflies Generations Genes Genetic Genetic Engineering Genome Genomics Glucose Goals Green Fluorescent Proteins Growth Harvest Homologous Gene Homoserine Human Immunologics In Vitro Infection Intravenous Iron Lead Leprosy Libraries Light Lipids Luciferases Metabolic Methionine Methods Modeling Mus Mycobacterium leprae Mycobacterium tuberculosis NADH Nude Mice Organ Organism Oxidoreductase Pantothenic Acid Pathway interactions Persons Pharmaceutical Preparations Pharmacotherapy Plasmids Predisposition Process Proteins Pseudogenes Radioactivity Recording of previous events Regimen Regulatory Pathway Reporter Reporting Research Research Personnel Research Technics Siderophores Signal Transduction Source Stains Stigmatization System Techniques Testing Therapeutic Agents Time Tissue imaging Tissues Work acid fast bacteria base cost drug development drug efficacy drug testing foot human disease improved in vivo imaging system intracellular parasitism mycobacterial mycobactins nanoluciferase novel novel therapeutics oxidation pathogen promoter tool uptake vaccine development

项目摘要

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.

项目概要/摘要麻风病在整个人类历史上一直困扰着人类，并且仍然困扰着 2 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它与人类疾病有关，但尚未在人工培养基上进行培养。它可以是在裸鼠的足垫中生长或在九带犰狳中全身生长。与一代人 14天的时间，通过计数抗酸杆菌来测试药物治疗方案，费力又费时。麻风分枝杆菌 DNA 序列与麻风分枝杆菌 DNA 序列的比较。结核病表明无法在人工培养基上培养麻风分枝杆菌可能是由于无法合成蛋氨酸、泛酸和分枝杆菌素。使用遗传工具，我们有为结核分枝杆菌开发，我们假设麻风分枝杆菌可以用噬菌体稳定转化基于整合熟练的质粒。我们计划采用生物发光共振能量转移（BRET）-Nanoluc和近红外蛋白进入质粒进行检测转化体。此外，我们假设麻风分枝杆菌的转化与缺失蛋氨酸和泛酸生物合成基因以及分枝杆菌素转运蛋白将使分枝杆菌成为可能。麻风杆菌在人工培养基上生长。一种发光的麻风分枝杆菌菌株可以增强药物效果发育和评估控制麻风分枝杆菌的免疫机制的能力感染。麻风分枝杆菌在人工培养基上生长的基因工程将增强我们的能力了解导致专性细胞内寄生的进化过程。