Development and Optimization of Chromosomally-Integrated Fluorescent Mycobacterium tuberculosis Reporter Constructs
Mycobacterium tuberculosis resides in the lungs in various lesion types with unique microenvironmental conditions. This diversity is in line with heterogeneous disease progression and divergent drug efficiency. Fluorescent reporter strains can be used to decipher the micromilieu and to guide future...
| Main Authors: | , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2020-12-01
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| Series: | Frontiers in Microbiology |
| Subjects: | |
| Online Access: | https://www.frontiersin.org/articles/10.3389/fmicb.2020.591866/full |
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| author | Katharina Kolbe Alice C. Bell Gareth A. Prosser Gareth A. Prosser Maike Assmann Hee-Jeong Yang He Eun Forbes Sophia Gallucci Katrin D. Mayer-Barber Helena I. Boshoff Clifton E. Barry III |
| author_facet | Katharina Kolbe Alice C. Bell Gareth A. Prosser Gareth A. Prosser Maike Assmann Hee-Jeong Yang He Eun Forbes Sophia Gallucci Katrin D. Mayer-Barber Helena I. Boshoff Clifton E. Barry III |
| author_sort | Katharina Kolbe |
| collection | DOAJ |
| description | Mycobacterium tuberculosis resides in the lungs in various lesion types with unique microenvironmental conditions. This diversity is in line with heterogeneous disease progression and divergent drug efficiency. Fluorescent reporter strains can be used to decipher the micromilieu and to guide future treatment regimens. Current reporters using replicating plasmids, however, are not suitable for long-term mouse infections or studies in non-human primates. Using a combination of recombinant DNA and protein optimization techniques, we have developed reporter strains based on integrative plasmids, which exhibit stimulus-response characteristics and fluorescence intensities comparable to those based on replicating plasmids. We successfully applied the concepts by constructing a multi-color reporter strain able to detect simultaneous changes in environmental pH, Mg2+ concentrations, and protein expression levels. |
| first_indexed | 2024-12-13T18:44:24Z |
| format | Article |
| id | doaj.art-eb56db4488294a2fb18e6b54d5fa9078 |
| institution | Directory Open Access Journal |
| issn | 1664-302X |
| language | English |
| last_indexed | 2024-12-13T18:44:24Z |
| publishDate | 2020-12-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Microbiology |
| spelling | doaj.art-eb56db4488294a2fb18e6b54d5fa90782022-12-21T23:35:07ZengFrontiers Media S.A.Frontiers in Microbiology1664-302X2020-12-011110.3389/fmicb.2020.591866591866Development and Optimization of Chromosomally-Integrated Fluorescent Mycobacterium tuberculosis Reporter ConstructsKatharina Kolbe0Alice C. Bell1Gareth A. Prosser2Gareth A. Prosser3Maike Assmann4Hee-Jeong Yang5He Eun Forbes6Sophia Gallucci7Katrin D. Mayer-Barber8Helena I. Boshoff9Clifton E. Barry III10Tuberculosis Research Section, Laboratory of Clinical Immunology and Microbiology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, United StatesTuberculosis Research Section, Laboratory of Clinical Immunology and Microbiology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, United StatesTuberculosis Research Section, Laboratory of Clinical Immunology and Microbiology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, United StatesDrug Discovery Unit, College of Life Sciences, James Black Centre, University of Dundee, Dundee, United KingdomInflammation and Innate Immunity Unit, Laboratory of Clinical Immunology and Microbiology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, United StatesTuberculosis Research Section, Laboratory of Clinical Immunology and Microbiology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, United StatesTuberculosis Research Section, Laboratory of Clinical Immunology and Microbiology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, United StatesTuberculosis Research Section, Laboratory of Clinical Immunology and Microbiology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, United StatesInflammation and Innate Immunity Unit, Laboratory of Clinical Immunology and Microbiology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, United StatesTuberculosis Research Section, Laboratory of Clinical Immunology and Microbiology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, United StatesTuberculosis Research Section, Laboratory of Clinical Immunology and Microbiology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, United StatesMycobacterium tuberculosis resides in the lungs in various lesion types with unique microenvironmental conditions. This diversity is in line with heterogeneous disease progression and divergent drug efficiency. Fluorescent reporter strains can be used to decipher the micromilieu and to guide future treatment regimens. Current reporters using replicating plasmids, however, are not suitable for long-term mouse infections or studies in non-human primates. Using a combination of recombinant DNA and protein optimization techniques, we have developed reporter strains based on integrative plasmids, which exhibit stimulus-response characteristics and fluorescence intensities comparable to those based on replicating plasmids. We successfully applied the concepts by constructing a multi-color reporter strain able to detect simultaneous changes in environmental pH, Mg2+ concentrations, and protein expression levels.https://www.frontiersin.org/articles/10.3389/fmicb.2020.591866/fullMycobacterium tuberculosisfluorescent proteinreporter strainriboswitchpHmagnesium |
| spellingShingle | Katharina Kolbe Alice C. Bell Gareth A. Prosser Gareth A. Prosser Maike Assmann Hee-Jeong Yang He Eun Forbes Sophia Gallucci Katrin D. Mayer-Barber Helena I. Boshoff Clifton E. Barry III Development and Optimization of Chromosomally-Integrated Fluorescent Mycobacterium tuberculosis Reporter Constructs Frontiers in Microbiology Mycobacterium tuberculosis fluorescent protein reporter strain riboswitch pH magnesium |
| title | Development and Optimization of Chromosomally-Integrated Fluorescent Mycobacterium tuberculosis Reporter Constructs |
| title_full | Development and Optimization of Chromosomally-Integrated Fluorescent Mycobacterium tuberculosis Reporter Constructs |
| title_fullStr | Development and Optimization of Chromosomally-Integrated Fluorescent Mycobacterium tuberculosis Reporter Constructs |
| title_full_unstemmed | Development and Optimization of Chromosomally-Integrated Fluorescent Mycobacterium tuberculosis Reporter Constructs |
| title_short | Development and Optimization of Chromosomally-Integrated Fluorescent Mycobacterium tuberculosis Reporter Constructs |
| title_sort | development and optimization of chromosomally integrated fluorescent mycobacterium tuberculosis reporter constructs |
| topic | Mycobacterium tuberculosis fluorescent protein reporter strain riboswitch pH magnesium |
| url | https://www.frontiersin.org/articles/10.3389/fmicb.2020.591866/full |
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