Miniaturization of Smart-seq2 for Single-Cell and Single-Nucleus RNA Sequencing

This protocol presents a plate-based workflow to perform RNA sequencing analysis of single cells/nuclei using Smart-seq2. We describe (1) the dissociation procedures for cell/nucleus isolation from the mouse brain and human organoids, (2) the flow sorting of single cells/nuclei into 384-well plates,...

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Autores principales: Jaeger, Baptiste N., Yángüez, Emilio, Gesuita, Lorenzo, Denoth-Lippuner, Annina, Kruse, Merit, Karayannis, Theofanis, Jessberger, Sebastian
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Elsevier 2020
Materias:
Protocol
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7501729/
https://www.ncbi.nlm.nih.gov/pubmed/33000004
http://dx.doi.org/10.1016/j.xpro.2020.100081
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author Jaeger, Baptiste N.
Yángüez, Emilio
Gesuita, Lorenzo
Denoth-Lippuner, Annina
Kruse, Merit
Karayannis, Theofanis
Jessberger, Sebastian
author_facet Jaeger, Baptiste N.
Yángüez, Emilio
Gesuita, Lorenzo
Denoth-Lippuner, Annina
Kruse, Merit
Karayannis, Theofanis
Jessberger, Sebastian
author_sort Jaeger, Baptiste N.
collection PubMed
description This protocol presents a plate-based workflow to perform RNA sequencing analysis of single cells/nuclei using Smart-seq2. We describe (1) the dissociation procedures for cell/nucleus isolation from the mouse brain and human organoids, (2) the flow sorting of single cells/nuclei into 384-well plates, and (3) the preparation of libraries following miniaturization of the Smart-seq2 protocol using a liquid-handling robot. This pipeline allows for the reliable, high-throughput, and cost-effective preparation of mouse and human samples for full-length deep single-cell/nucleus RNA sequencing. For complete details on the use and execution of this protocol, please refer to Bowers et al. (2020).
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spelling pubmed-75017292020-09-28 Miniaturization of Smart-seq2 for Single-Cell and Single-Nucleus RNA Sequencing Jaeger, Baptiste N. Yángüez, Emilio Gesuita, Lorenzo Denoth-Lippuner, Annina Kruse, Merit Karayannis, Theofanis Jessberger, Sebastian STAR Protoc Protocol This protocol presents a plate-based workflow to perform RNA sequencing analysis of single cells/nuclei using Smart-seq2. We describe (1) the dissociation procedures for cell/nucleus isolation from the mouse brain and human organoids, (2) the flow sorting of single cells/nuclei into 384-well plates, and (3) the preparation of libraries following miniaturization of the Smart-seq2 protocol using a liquid-handling robot. This pipeline allows for the reliable, high-throughput, and cost-effective preparation of mouse and human samples for full-length deep single-cell/nucleus RNA sequencing. For complete details on the use and execution of this protocol, please refer to Bowers et al. (2020). Elsevier 2020-08-05 /pmc/articles/PMC7501729/ /pubmed/33000004 http://dx.doi.org/10.1016/j.xpro.2020.100081 Text en © 2020 The Authors 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
Jaeger, Baptiste N.
Yángüez, Emilio
Gesuita, Lorenzo
Denoth-Lippuner, Annina
Kruse, Merit
Karayannis, Theofanis
Jessberger, Sebastian
Miniaturization of Smart-seq2 for Single-Cell and Single-Nucleus RNA Sequencing
title Miniaturization of Smart-seq2 for Single-Cell and Single-Nucleus RNA Sequencing
title_full Miniaturization of Smart-seq2 for Single-Cell and Single-Nucleus RNA Sequencing
title_fullStr Miniaturization of Smart-seq2 for Single-Cell and Single-Nucleus RNA Sequencing
title_full_unstemmed Miniaturization of Smart-seq2 for Single-Cell and Single-Nucleus RNA Sequencing
title_short Miniaturization of Smart-seq2 for Single-Cell and Single-Nucleus RNA Sequencing
title_sort miniaturization of smart-seq2 for single-cell and single-nucleus rna sequencing
topic Protocol
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7501729/
https://www.ncbi.nlm.nih.gov/pubmed/33000004
http://dx.doi.org/10.1016/j.xpro.2020.100081
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