FACS-Based Neuronal Cell Type-Specific RNA Isolation and Alternative Splicing Analysis.
Cell type–specific
FACS
Pre-mRNA splicing
Journal
Methods in molecular biology (Clifton, N.J.)
ISSN: 1940-6029
Titre abrégé: Methods Mol Biol
Pays: United States
ID NLM: 9214969
Informations de publication
Date de publication:
2022
2022
Historique:
entrez:
27
7
2022
pubmed:
28
7
2022
medline:
30
7
2022
Statut:
ppublish
Résumé
Alternative splicing of pre-mRNAs expands the coding abilities of genomes by generating distinct transcription variants from individual genes. It contributes to the marvelous complexity of the transcriptome in neurons. Given the differential expression of alternative splicing regulators and diversity in alternative splicing programs in neuronal subpopulations, it is urgent and necessary to develop methods to efficiently isolate diverse subgroups of neurons and analyze their transcriptomic diversity. Here, we describe a protocol to isolate RNA from specific neuronal types using a fluorescence-activated cell sorting (FACS)-based method to analyze alternative splicing events in a cell type-specific manner. The method is universally applicable to analyze alternative splicing in fluorescent protein-labeled neuronal types. It was optimized to preserve the transcription state and improve efficiency in cell suspension purification. With our protocol, fluorescent protein-labeled neurons could be efficiently purified. The transcription states suitable for gene expression and alternative splicing analysis could be well-preserved.
Identifiants
pubmed: 35895258
doi: 10.1007/978-1-0716-2521-7_4
doi:
Substances chimiques
RNA Precursors
0
RNA
63231-63-0
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
51-62Informations de copyright
© 2022. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.
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