رسالة جامعية
Achieving Single-Molecule Tracking in Bacteria During Real-Time Environmental Perturbations
| العنوان: | Achieving Single-Molecule Tracking in Bacteria During Real-Time Environmental Perturbations |
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| المؤلفون: | Calkins, Anna |
| المساهمون: | Biteen, Julie Suzanne, Sandkvist, Maria B, Bailey, Ryan Castle, Kennedy, Robert T |
| سنة النشر: | 2023 |
| المجموعة: | University of Michigan: Deep Blue |
| مصطلحات موضوعية: | Single-molecule Fluorescence Microscopy, Microfluidics, Real-time Environment Perturbations, Live Bacteria Cell Imaging, Bacteria Cell Sample Preparation Methods, Chemistry, Science |
| الوصف: | Bacteria have developed a variety of mechanisms to adapt to changes in the environment in ways that are often vital to their proliferation and survival. Single-molecule fluorescence imaging and tracking have elucidated the complex mechanisms of these responses in a variety of bacteria. However, a technological gap remains: due to cell sample preparation limitations, all of the SMF experiments in live bacteria are performed at steady state such that all components have reached equilibrium. Therefore, this dissertation aims to address this gap in technology and access single-molecule dynamics during real-time changes by implementing two alternative sample preparations: microfluidic devices and chitosan-coated coverslips. Chapter I discusses bacteria adaptation to environment changes and detailed information regarding one example discussed throughout this dissertation, V. cholerae. It explores the optics principles of single-molecule imaging and how it improves the localization precision of optical imaging. As an example of diffraction-limited single-molecule imaging, Chapter II discusses imaging Colicin E1 in live E. coli cells. Finally, the key limitation of SMF in live bacteria cells is discussed. V. cholerae uses a membrane-localized transcription factor, TcpP, to regulate toxin production in response to changes in the environment. Chapter III describes SMF imaging and tracking performed on TcpP-PAmCherry in live V. cholerae cells. The experiments discussed in Chapter III determined that TcpP and ToxR work cooperatively under steady-state conditions, but measurements of how these dynamical interactions change over the course of environmental perturbations were precluded by the traditional preparation of bacterial cells confined on agarose pads. This challenge leads to Chapter IV, which describes the first novel sample preparation method to address this gap: chitosan-coated coverslips. The chapter discusses the procedure of making chitosan-coated coverslips, confirms the suitability of these coverslips for ... |
| نوع الوثيقة: | thesis |
| وصف الملف: | application/pdf |
| اللغة: | English |
| العلاقة: | https://hdl.handle.net/2027.42/176624Test; https://dx.doi.org/10.7302/7473Test; orcid:0000-0003-0130-296X; Calkins, Anna; 0000-0003-0130-296X |
| DOI: | 10.7302/7473 |
| الإتاحة: | https://doi.org/10.7302/7473Test https://hdl.handle.net/2027.42/176624Test |
| رقم الانضمام: | edsbas.1F14C84C |
| قاعدة البيانات: | BASE |
| DOI: | 10.7302/7473 |
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