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Direct isolation of single cells from living brains of Drosophila melanogaster without dissociation for transcriptome analysis

Here, we describe a protocol to remove single identified cells directly from Drosophila living brains and analyze their transcriptome. We detail the steps to harvest fluorescent cells using a capillary under epifluorescence and transmitted light to avoid contamination. We then outline the procedure...

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Detalles Bibliográficos
Autores principales: Barros, Claudia S., Bossing, Torsten
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Elsevier 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9529598/
https://www.ncbi.nlm.nih.gov/pubmed/36181682
http://dx.doi.org/10.1016/j.xpro.2022.101735
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author Barros, Claudia S.
Bossing, Torsten
author_facet Barros, Claudia S.
Bossing, Torsten
author_sort Barros, Claudia S.
collection PubMed
description Here, we describe a protocol to remove single identified cells directly from Drosophila living brains and analyze their transcriptome. We detail the steps to harvest fluorescent cells using a capillary under epifluorescence and transmitted light to avoid contamination. We then outline the procedure to obtain the transcriptome by reverse transcription and amplification. The process from cell harvesting to the initiation of reverse transcription only takes 2 min, thus avoiding transcriptional activation of cell damage response or cell death genes. For complete details on the use and execution of this protocol, please refer to Barros and Bossing (2021), Bossing et al. (2012), Gil-Ranedo et al. (2019), and Liu and Bossing (2016).
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spelling pubmed-95295982022-10-05 Direct isolation of single cells from living brains of Drosophila melanogaster without dissociation for transcriptome analysis Barros, Claudia S. Bossing, Torsten STAR Protoc Protocol Here, we describe a protocol to remove single identified cells directly from Drosophila living brains and analyze their transcriptome. We detail the steps to harvest fluorescent cells using a capillary under epifluorescence and transmitted light to avoid contamination. We then outline the procedure to obtain the transcriptome by reverse transcription and amplification. The process from cell harvesting to the initiation of reverse transcription only takes 2 min, thus avoiding transcriptional activation of cell damage response or cell death genes. For complete details on the use and execution of this protocol, please refer to Barros and Bossing (2021), Bossing et al. (2012), Gil-Ranedo et al. (2019), and Liu and Bossing (2016). Elsevier 2022-09-30 /pmc/articles/PMC9529598/ /pubmed/36181682 http://dx.doi.org/10.1016/j.xpro.2022.101735 Text en © 2022 The Author(s) https://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
Barros, Claudia S.
Bossing, Torsten
Direct isolation of single cells from living brains of Drosophila melanogaster without dissociation for transcriptome analysis
title Direct isolation of single cells from living brains of Drosophila melanogaster without dissociation for transcriptome analysis
title_full Direct isolation of single cells from living brains of Drosophila melanogaster without dissociation for transcriptome analysis
title_fullStr Direct isolation of single cells from living brains of Drosophila melanogaster without dissociation for transcriptome analysis
title_full_unstemmed Direct isolation of single cells from living brains of Drosophila melanogaster without dissociation for transcriptome analysis
title_short Direct isolation of single cells from living brains of Drosophila melanogaster without dissociation for transcriptome analysis
title_sort direct isolation of single cells from living brains of drosophila melanogaster without dissociation for transcriptome analysis
topic Protocol
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9529598/
https://www.ncbi.nlm.nih.gov/pubmed/36181682
http://dx.doi.org/10.1016/j.xpro.2022.101735
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