Isolation of Mitochondria from Liver and Extraction of Total RNA and Protein: Analyses of miRNA and Protein Expression

André L. Simão, Pedro M. Borralho, Clifford J. Steer, Rui E. Castro, Cecília M.P. Rodrigues

Research output: Chapter in Book/Report/Conference proceedingChapter

1 Scopus citations

Abstract

Several studies have indicated the presence of microRNAs (miRNAs) within mitochondria although the origin, as well as the biological function, of these mitochondrially located miRNAs is largely unknown. The identification and significance of this subcellular localization is gaining increasing relevance to the pathogenesis of certain disease states. Here, we describe the isolation of highly purified mitochondria from rat liver by differential centrifugation, followed by RNAse A treatment to eliminate contaminating RNA. The coupled extraction of total RNA and protein is a more efficient design for allowing the downstream evaluation of miRNA and protein expression in mitochondria.

Original languageEnglish (US)
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc.
Pages1-15
Number of pages15
Volume2310
DOIs
StatePublished - 2021

Publication series

NameMethods in Molecular Biology
Volume2310
ISSN (Print)1064-3745
ISSN (Electronic)1940-6029

Bibliographical note

Funding Information:
CMPR is supported by the EU H2020 Marie Sklodowska-Curie Project Foie Gras (grant 722619) and by FCT and FEDER (grant PTDC/MEDFAR/29097/2017 e LISBOA-01-0145-FEDER-029097).

Publisher Copyright:
© 2021, Springer Science+Business Media, LLC, part of Springer Nature.

Keywords

  • Differential centrifugation
  • Mitochondria
  • Protein
  • TRIzol™
  • miRNA
  • Cell Fractionation
  • Mitochondrial Proteins/isolation & purification
  • Rats
  • Ultracentrifugation
  • MicroRNAs/isolation & purification
  • Ribonuclease, Pancreatic/metabolism
  • Mitochondria, Liver/metabolism
  • Animals
  • Real-Time Polymerase Chain Reaction

PubMed: MeSH publication types

  • Research Support, Non-U.S. Gov't
  • Journal Article

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