Cell Type–Specific Isolation of Mitochondria in Arabidopsis. Boussardon, C. & Keech, O. In Van Aken, O. & Rasmusson, A. G., editors, Plant Mitochondria: Methods and Protocols, of Methods in Molecular Biology, pages 13–23. Springer US, New York, NY, January, 2022. ![Cell Type–Specific Isolation of Mitochondria in Arabidopsis [link]](https://bibbase.org/img/filetypes/link.svg "link")
Paper abstract bibtex Membrane-bound organelles are unique features of eukaryotic cell structures. Among them, mitochondria host key metabolic functions and pathways, including the aerobic respiration. In plants, several procedures are available to isolate mitochondria from the other cell compartments, as high-quality purified extracts are often necessary for accurate molecular biology or biochemistry investigations. Protocols based on differential centrifugations and subsequent density gradients are an effective way to extract rather pure and intact mitochondria within a few hours. However, while mitochondria from seedlings, large leaves or tubers are relatively easy to extract, tissue-specific isolation of organelles had remained a challenge. This has recently been circumvented, only in transformable plants though, by the use of affinity-tagged mitochondria and their isolation with magnetic beads.We hereby describe a step-by-step protocol for the rapid and tissue-specific isolation of Arabidopsis thaliana mitochondria, a method named IMTACT (Isolation of Mitochondria TAgged in specific Cell Types). Cell-specific biotinylated mitochondria are isolated with streptavidin magnetic beads in less than 30 min from sampling to final extract. Key steps, enrichment, bead size comparison, and mitochondrial depletion in the sample are also reported in order to facilitate the experimental setup of the user.
@incollection{boussardon_cell_2022,
address = {New York, NY},
series = {Methods in {Molecular} {Biology}},
title = {Cell {Type}–{Specific} {Isolation} of {Mitochondria} in {Arabidopsis}},
isbn = {978-1-07-161653-6},
url = {https://doi.org/10.1007/978-1-0716-1653-6_2},
abstract = {Membrane-bound organelles are unique features of eukaryotic cell structures. Among them, mitochondria host key metabolic functions and pathways, including the aerobic respiration. In plants, several procedures are available to isolate mitochondria from the other cell compartments, as high-quality purified extracts are often necessary for accurate molecular biology or biochemistry investigations. Protocols based on differential centrifugations and subsequent density gradients are an effective way to extract rather pure and intact mitochondria within a few hours. However, while mitochondria from seedlings, large leaves or tubers are relatively easy to extract, tissue-specific isolation of organelles had remained a challenge. This has recently been circumvented, only in transformable plants though, by the use of affinity-tagged mitochondria and their isolation with magnetic beads.We hereby describe a step-by-step protocol for the rapid and tissue-specific isolation of Arabidopsis thaliana mitochondria, a method named IMTACT (Isolation of Mitochondria TAgged in specific Cell Types). Cell-specific biotinylated mitochondria are isolated with streptavidin magnetic beads in less than 30 min from sampling to final extract. Key steps, enrichment, bead size comparison, and mitochondrial depletion in the sample are also reported in order to facilitate the experimental setup of the user.},
language = {en},
urldate = {2021-09-23},
booktitle = {Plant {Mitochondria}: {Methods} and {Protocols}},
publisher = {Springer US},
author = {Boussardon, Clément and Keech, Olivier},
editor = {Van Aken, Olivier and Rasmusson, Allan G.},
month = jan,
year = {2022},
keywords = {Biotin–streptavidin interaction, Editable Golden Gate plasmids, Mitochondria, Tagged outer membrane, Tissue-specific isolation},
pages = {13--23},
}
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Protocols based on differential centrifugations and subsequent density gradients are an effective way to extract rather pure and intact mitochondria within a few hours. However, while mitochondria from seedlings, large leaves or tubers are relatively easy to extract, tissue-specific isolation of organelles had remained a challenge. This has recently been circumvented, only in transformable plants though, by the use of affinity-tagged mitochondria and their isolation with magnetic beads.We hereby describe a step-by-step protocol for the rapid and tissue-specific isolation of Arabidopsis thaliana mitochondria, a method named IMTACT (Isolation of Mitochondria TAgged in specific Cell Types). Cell-specific biotinylated mitochondria are isolated with streptavidin magnetic beads in less than 30 min from sampling to final extract. 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