Abstract
The overexpression of a specific protein is a common method for investigating the specific biological function of the substance and the mechanism of action. In vivo electrotransfer has been confirmed to be one of the most reliable, efficient and cost-effective way to overexpress a protein in a select biological tissue. Typically, this technique involves a physical injection of plasmid DNA followed by electric pulses across the injection site. Here, we introduce this method that we used to transfect green fluorescent protein (GFP)-tagged PGC-1α plasmid DNA into mouse tibialis anterior (TA) muscle, which attained high transfection efficiency with no muscle damage. To quantify the transfection efficiency, we also demonstrate the visualization of plasmid DNA transfected fibers via immunohistochemical staining on muscle cross sections.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Wolff JA, Ludtke JJ, Acsadi G et al (1992) Long-term persistence of plasmid DNA and foreign gene expression in mouse muscle. Hum Mol Genet 1:363–369
Davis HL, Demeneix BA, Quantin B et al (1993) Plasmid DNA is superior to viral vectors for direct gene transfer into adult mouse skeletal muscle. Hum Gene Ther 4:733–740. https://doi.org/10.1089/hum.1993.4.6-733
Trollet C, Bloquel C, Scherman D, Bigey P (2006) Electrotransfer into skeletal muscle for protein expression. Curr Gene Ther 6:561–578
Mir LM, Bureau MF, Gehl J et al (1999) High-efficiency gene transfer into skeletal muscle mediated by electric pulses. Proc Natl Acad Sci U S A 96:4262–4267
Aihara H, Miyazaki J (1998) Gene transfer into muscle by electroporation in vivo. Nat Biotechnol 16:867–870. https://doi.org/10.1038/nbt0998-867
Cheng L, Ziegelhoffer PR, Yang NS (1993) In vivo promoter activity and transgene expression in mammalian somatic tissues evaluated by using particle bombardment. Proc Natl Acad Sci U S A 90:4455–4459
Heller LC, Ugen K, Heller R (2005) Electroporation for targeted gene transfer. Expert Opin Drug Deliv 2:255–268. https://doi.org/10.1517/17425247.2.2.255
McMahon JM, Wells DJ (2004) Electroporation for gene transfer to skeletal muscles: current status. BioDrugs 18:155–165
Nayerossadat N, Maedeh T, Ali PA (2012) Viral and nonviral delivery systems for gene delivery. Adv Biomed Res 1:27. https://doi.org/10.4103/2277-9175.98152
Schertzer JD, Plant DR, Lynch GS (2006) Optimizing plasmid-based gene transfer for investigating skeletal muscle structure and function. Mol Ther 13:795–803. https://doi.org/10.1016/j.ymthe.2005.09.019
Taylor J, Babbs CF, Alzghoul MB et al (2004) Optimization of ectopic gene expression in skeletal muscle through DNA transfer by electroporation. BMC Biotechnol 4:11–11. https://doi.org/10.1186/1472-6750-4-11
Donà M, Sandri M, Rossini K et al (2003) Functional in vivo gene transfer into the myofibers of adult skeletal muscle. Biochem Biophys Res Commun 312:1132–1138. https://doi.org/10.1016/j.bbrc.2003.11.032
Kang C, Goodman CA, Hornberger TA, Ji LL (2015) PGC-1α overexpression by in vivo transfection attenuates mitochondrial deterioration of skeletal muscle caused by immobilization. FASEB J 29:4092–4106. https://doi.org/10.1096/fj.14-266619
Kang C, Ji LL (2016) PGC-1α overexpression via local transfection attenuates mitophagy pathway in muscle disuse atrophy. Free Radic Biol Med 93:32–40. https://doi.org/10.1016/j.freeradbiomed.2015.12.032
Puigserver P, Wu Z, Park CW et al (1998) A cold-inducible coactivator of nuclear receptors linked to adaptive thermogenesis. Cell 92:829–839
Taketo M, Schroeder AC, Mobraaten LE et al (1991) FVB/N: an inbred mouse strain preferable for transgenic analyses. PNAS 88:2065–2069. https://doi.org/10.1073/pnas.88.6.2065
Yeo D, Kang C, Gomez-Cabrera MC et al (2019) Intensified mitophagy in skeletal muscle with aging is downregulated by PGC-1alpha overexpression in vivo. Free Radic Biol Med 130:361–368. https://doi.org/10.1016/j.freeradbiomed.2018.10.456
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2019 Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Yeo, D., Kang, C., Ji, L.L. (2019). PGC-1α Overexpression via Local In Vivo Transfection in Mouse Skeletal Muscle. In: Badr, M. (eds) Nuclear Receptors. Methods in Molecular Biology, vol 1966. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9195-2_12
Download citation
DOI: https://doi.org/10.1007/978-1-4939-9195-2_12
Published:
Publisher Name: Humana, New York, NY
Print ISBN: 978-1-4939-9194-5
Online ISBN: 978-1-4939-9195-2
eBook Packages: Springer Protocols