Mitochondrial targeting of recombinant RNA.

Methods Mol Biol

UMR 7156 Genetique Moleculaire, Genomique, Microbiologie (GMGM), University of Strasbourg-CNRS, 21 Rue René Descartes, 67084, Strasbourg, France.

Published: October 2015

Mitochondrial import of small noncoding RNA is found in a large variety of species. In mammalian cells, this pathway can be used for therapeutic purpose, to restore the mitochondrial functions affected by pathogenic mutations. Recently, we developed mitochondrial RNA vectors able to address therapeutic oligoribonucleotides into human mitochondria. Here we provide the protocol for transfection of cultured human cells with small recombinant RNA molecules and describe two approaches useful to demonstrate their import into mitochondria: (1) isolation of RNA from purified mitochondria and quantitative hybridization analysis and (2) confocal microscopy of cells transfected with fluorescently labeled RNA. These protocols can be used in combination with overexpression or downregulation of protein import factors to detect and to evaluate their influence on the mitochondrial import of various RNAs.

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Source
http://dx.doi.org/10.1007/978-1-4939-2288-8_16DOI Listing

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