Chapter title |
In Situ Imaging of Bacterial Secretion Systems by Electron Cryotomography
|
---|---|
Chapter number | 27 |
Book title |
Bacterial Protein Secretion Systems
|
Published in |
Methods in molecular biology, July 2017
|
DOI | 10.1007/978-1-4939-7033-9_27 |
Pubmed ID | |
Book ISBNs |
978-1-4939-7031-5, 978-1-4939-7033-9
|
Authors |
Gregor L. Weiss, João M. Medeiros, Martin Pilhofer |
Editors |
Laure Journet, Eric Cascales |
Abstract |
The unique property of electron cryotomography (ECT) is its capability to resolve the structure of macromolecular machines in their cellular context. The integration of ECT data with high-resolution structures of purified subcomplexes and live-cell fluorescence light microscopy can generate pseudo-atomic models that lead to a mechanistic understanding across size and time scales. Recent advances in electron detection, sample thinning, data acquisition, and data processing have significantly enhanced the applicability and performance of ECT. Here we describe a detailed workflow for an ECT experiment, including cell culture, vitrification, data acquisition, data reconstruction, tomogram analysis, and subtomogram averaging. This protocol provides an entry point to the technique for students and researchers and indicates the many possible variations arising from specific target properties and the available instrumentation. |
Mendeley readers
Geographical breakdown
Country | Count | As % |
---|---|---|
Unknown | 36 | 100% |
Demographic breakdown
Readers by professional status | Count | As % |
---|---|---|
Student > Ph. D. Student | 10 | 28% |
Student > Doctoral Student | 6 | 17% |
Researcher | 4 | 11% |
Student > Bachelor | 3 | 8% |
Student > Postgraduate | 2 | 6% |
Other | 4 | 11% |
Unknown | 7 | 19% |
Readers by discipline | Count | As % |
---|---|---|
Biochemistry, Genetics and Molecular Biology | 13 | 36% |
Immunology and Microbiology | 7 | 19% |
Agricultural and Biological Sciences | 6 | 17% |
Computer Science | 1 | 3% |
Arts and Humanities | 1 | 3% |
Other | 2 | 6% |
Unknown | 6 | 17% |