Akhuli, D and Dhar, A and Viji, AS and Bhojappa, B and Palani, S (2022) ALIBY: ALFA Nanobody-Based Toolkit for Imaging and Biochemistry in Yeast. In: mSphere, 7 (5).
Full text not available from this repository.Abstract
Specialized epitope tags continue to be integral components of various biochemical and cell biological applications such as fluorescence microscopy, immunoblotting, immunoprecipitation, and protein purification. However, until recently, no single tag could offer this complete set of functionalities on its own. Here, we present a plasmid-based toolkit named ALIBY (ALFA toolkit for imaging and biochemistry in yeast) that provides a universal workflow to adopt the versatile ALFA tag/ NbALFA system within the well-established model organism Saccharomyces cerevisiae. The kit comprises tagging plasmids for labeling a protein of interest with the ALFA tag and detection plasmids encoding fluorescent-protein-tagged NbALFA for live-cell imaging purposes. We demonstrate the suitability of ALIBY for visualizing the spatiotemporal localization of yeast proteins (i.e., the cytoskeleton, nucleus, centrosome, mitochondria, vacuole, endoplasmic reticulum, exocyst, and divisome) in live cells. Our approach has yielded an excellent signal-to-noise ratio without off-target effects or any effect on cell growth. In summary, our yeast-specific toolkit aims to simplify and further advance the live-cell imaging of differentially abundant yeast proteins while also being suitable for biochemical applications. © 2022 Akhuli et al.
| Item Type: | Journal Article |
|---|---|
| Publication: | mSphere |
| Publisher: | American Society for Microbiology |
| Additional Information: | The copyright for this article belongs to the Authors. |
| Keywords: | epitope; Saccharomyces cerevisiae protein, chemistry; endoplasmic reticulum; genetics; metabolism; plasmid; Saccharomyces cerevisiae, Endoplasmic Reticulum; Epitopes; Plasmids; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins |
| Department/Centre: | Division of Biological Sciences > Biochemistry |
| Date Deposited: | 22 Nov 2022 05:08 |
| Last Modified: | 22 Nov 2022 05:08 |
| URI: | https://eprints.iisc.ac.in/id/eprint/77897 |
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