Super-resolution microscopy and single-protein tracking in live bacteria using a genetically encoded, photostable fluoromodule

Saumya Saurabh, Adam M. Perez, Colin J. Comerci, Lucy Shapiro, W. E. Moerner

Research output: Contribution to journalArticlepeer-review

Abstract

Visualization of dynamic protein structures in live cells is crucial for understanding the mechanisms governing biological processes. Fluorescence microscopy is a sensitive tool for this purpose. In order to image proteins in live bacteria using fluorescence microscopy, one typically genetically fuses the protein of interest to a photostable fluorescent tag. Several labeling schemes are available to accomplish this. Particularly, hybrid tags that combine a fluorescent or fluorogenic dye with a genetically encoded protein (such as enzymatic labels) have been used successfully in multiple cell types. However, their use in bacteria has been limited due to challenges imposed by a complex bacterial cell wall. Here, we describe the use of a genetically encoded photostable fluoromodule that can be targeted to cytosolic and membrane proteins in the Gram negative bacterium Caulobacter crescentus. Additionally, we summarize methods to use this fluoromodule for single protein imaging and super-resolution microscopy using stimulated emission depletion.

Original languageEnglish (US)
Pages (from-to)4.32.1-4.32.22
JournalCurrent Protocols in Cell Biology
Volume2017
DOIs
StatePublished - Jun 1 2017

Keywords

  • Bacteria
  • Fluorogenic
  • Fluoromodule
  • Photostable
  • Single-protein tracking
  • STED

ASJC Scopus subject areas

  • Cell Biology

Fingerprint

Dive into the research topics of 'Super-resolution microscopy and single-protein tracking in live bacteria using a genetically encoded, photostable fluoromodule'. Together they form a unique fingerprint.

Cite this