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Protocol for Proximity-Dependent Proteomic Profiling in Yeast Cells by APEX and Alk-Ph Probe

Alk-Ph is a clickable APEX2 substrate developed for spatially restricted protein/RNA labeling in intact yeast cells. Alk-Ph is more water soluble and cell wall permeable than biotin-phenol substrate, allowing more efficient profiling of the subcellular proteome in microorganisms. We describe the pro...

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Detalles Bibliográficos
Autores principales: Li, Yi, Liu, Keke, Zhou, Ying, Yang, Jing, Zou, Peng
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Elsevier 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7757286/
https://www.ncbi.nlm.nih.gov/pubmed/33377031
http://dx.doi.org/10.1016/j.xpro.2020.100137
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author Li, Yi
Liu, Keke
Zhou, Ying
Yang, Jing
Zou, Peng
author_facet Li, Yi
Liu, Keke
Zhou, Ying
Yang, Jing
Zou, Peng
author_sort Li, Yi
collection PubMed
description Alk-Ph is a clickable APEX2 substrate developed for spatially restricted protein/RNA labeling in intact yeast cells. Alk-Ph is more water soluble and cell wall permeable than biotin-phenol substrate, allowing more efficient profiling of the subcellular proteome in microorganisms. We describe the protocol for Alk-Ph probe synthesis, APEX2 expression, and protein/RNA labeling in yeast and the workflow for quantitative proteomic experiments and data analysis. Using the yeast mitochondria as an example, we provide guidelines to achieve high-resolution mapping of subcellular yeast proteome and transcriptome. For complete details on the use and execution of this protocol, please refer to Li et al. (2020).
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spelling pubmed-77572862020-12-28 Protocol for Proximity-Dependent Proteomic Profiling in Yeast Cells by APEX and Alk-Ph Probe Li, Yi Liu, Keke Zhou, Ying Yang, Jing Zou, Peng STAR Protoc Protocol Alk-Ph is a clickable APEX2 substrate developed for spatially restricted protein/RNA labeling in intact yeast cells. Alk-Ph is more water soluble and cell wall permeable than biotin-phenol substrate, allowing more efficient profiling of the subcellular proteome in microorganisms. We describe the protocol for Alk-Ph probe synthesis, APEX2 expression, and protein/RNA labeling in yeast and the workflow for quantitative proteomic experiments and data analysis. Using the yeast mitochondria as an example, we provide guidelines to achieve high-resolution mapping of subcellular yeast proteome and transcriptome. For complete details on the use and execution of this protocol, please refer to Li et al. (2020). Elsevier 2020-10-24 /pmc/articles/PMC7757286/ /pubmed/33377031 http://dx.doi.org/10.1016/j.xpro.2020.100137 Text en © 2020 The Author(s) http://creativecommons.org/licenses/by-nc-nd/4.0/ This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
spellingShingle Protocol
Li, Yi
Liu, Keke
Zhou, Ying
Yang, Jing
Zou, Peng
Protocol for Proximity-Dependent Proteomic Profiling in Yeast Cells by APEX and Alk-Ph Probe
title Protocol for Proximity-Dependent Proteomic Profiling in Yeast Cells by APEX and Alk-Ph Probe
title_full Protocol for Proximity-Dependent Proteomic Profiling in Yeast Cells by APEX and Alk-Ph Probe
title_fullStr Protocol for Proximity-Dependent Proteomic Profiling in Yeast Cells by APEX and Alk-Ph Probe
title_full_unstemmed Protocol for Proximity-Dependent Proteomic Profiling in Yeast Cells by APEX and Alk-Ph Probe
title_short Protocol for Proximity-Dependent Proteomic Profiling in Yeast Cells by APEX and Alk-Ph Probe
title_sort protocol for proximity-dependent proteomic profiling in yeast cells by apex and alk-ph probe
topic Protocol
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7757286/
https://www.ncbi.nlm.nih.gov/pubmed/33377031
http://dx.doi.org/10.1016/j.xpro.2020.100137
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