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Efficient transfection of DNA or shRNA vectors into neurons using magnetofection


Buerli, T; Pellegrino, C; Baer, K; Lardi-Studler, B; Chudotvorova, I; Fritschy, J M; Medina, I; Fuhrer, C (2007). Efficient transfection of DNA or shRNA vectors into neurons using magnetofection. Nature Protocols, 2(12):3090-3101.

Abstract

Efficient and long-lasting transfection of primary neurons is an essential tool to address many questions in current neuroscience using functional gene analysis. Neurons are sensitive to cytotoxicity and difficult to transfect with most methods. We provide a protocol for transfection of cDNA and RNA interference (shRNA) vectors, using magnetofection, into rat hippocampal neurons (E18/19) cultured for several hours to 21 days in vitro. This protocol even allows for double-transfection of DNA into a small subpopulation of hippocampal neurons (GABAergic interneurons), as well as achieving long-lasting expression of DNA and shRNA constructs without interfering with neuronal differentiation. The protocol, which uses inexpensive equipment and reagents, takes 1 h, utilizes mixed hippocampal cultures, a transfection reagent, CombiMag and a magnetic plate, shows low toxicity and is suited for single cell analysis. Modifications as done by our three laboratories are detailed.

Abstract

Efficient and long-lasting transfection of primary neurons is an essential tool to address many questions in current neuroscience using functional gene analysis. Neurons are sensitive to cytotoxicity and difficult to transfect with most methods. We provide a protocol for transfection of cDNA and RNA interference (shRNA) vectors, using magnetofection, into rat hippocampal neurons (E18/19) cultured for several hours to 21 days in vitro. This protocol even allows for double-transfection of DNA into a small subpopulation of hippocampal neurons (GABAergic interneurons), as well as achieving long-lasting expression of DNA and shRNA constructs without interfering with neuronal differentiation. The protocol, which uses inexpensive equipment and reagents, takes 1 h, utilizes mixed hippocampal cultures, a transfection reagent, CombiMag and a magnetic plate, shows low toxicity and is suited for single cell analysis. Modifications as done by our three laboratories are detailed.

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Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:04 Faculty of Medicine > Brain Research Institute
Dewey Decimal Classification:570 Life sciences; biology
610 Medicine & health
Uncontrolled Keywords:Magnetofection, Transfection, Protein Expression, Neuronal cultures, Hippocampus
Language:English
Date:2007
Deposited On:11 Feb 2008 12:13
Last Modified:05 Apr 2016 12:12
Publisher:Nature Publishing Group
ISSN:1750-2799
Publisher DOI:https://doi.org/10.1038/nprot.2007.445
PubMed ID:18079708

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