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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...
Autores principales: | , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Elsevier
2020
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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). |
format | Online Article Text |
id | pubmed-7757286 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | Elsevier |
record_format | MEDLINE/PubMed |
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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