Skip to main content

Complexation of siRNA and pDNA with Cationic Liposomes: The Important Aspects in Lipoplex Preparation

  • Protocol
  • First Online:
Liposomes

Part of the book series: Methods in Molecular Biology ((MIMB,volume 605))

Abstract

In the last two decades, cationic liposomes have been investigated as vehicles for nucleic acids [plasmid DNA (pDNA) and small interfering RNA (siRNA)] delivery in vitro and in vivo. The formation of cationic liposomes–nucleic acids complexes, termed lipoplexes, depends on a number of experimental variables. The quality of the nucleic acid and the cationic liposome as well as the selection of diluents for diluting the concentrated stocks strongly affect the resulting lipoplexes and their efficiency of gene-expression or gene-silencing effect following transfection. In addition, the molar ratio of cationic lipid nitrogen (N) to siRNA or pDNA phosphate (P) (N/P ratio) influences the final characteristics of the lipoplexes, such as size, surface zeta potential, and reproducibility, thereby reflecting their efficiency following transfection. The methods presented in this chapter could be helpful to obtain reliable and reproducible lipoplexes and experimental results.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 89.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 119.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 169.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Templeton NS, Lasic DD, Frederik PM, Strey HH, Roberts DD, Pavlakis GN (1997) Improved DNA: liposome complexes for increased systemic delivery and gene expression. Nat Biotechnol 15:647–652

    Article  CAS  PubMed  Google Scholar 

  2. Elbashir SM, Harborth J, Lendeckel W, Yalcin A, Weber K, Tuschl T (2001) Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells. Nature 411:494–498

    Article  CAS  PubMed  Google Scholar 

  3. Dass CR, Choong PF (2006) Selective gene delivery for cancer therapy using cationic liposomes: in vivo proof of applicability. J Control Release 113:155–163

    Article  CAS  PubMed  Google Scholar 

  4. Zhang S, Zhao B, Jiang H, Wang B, Ma B (2007) Cationic lipids and polymers mediated vectors for delivery of siRNA. J Control Release 123:1–10

    Article  CAS  PubMed  Google Scholar 

  5. Garcia-Chaumont C, Seksek O, Grzybowska J, Borowski E, Bolard J (2000) Delivery systems for antisense oligonucleotides. Pharmacol Ther 87:255–277

    Article  CAS  PubMed  Google Scholar 

  6. Lasic DD, Templeton NS (1996) Liposome in gene therapy. Adv Drug Deliv Rev 20:221–266

    Article  CAS  Google Scholar 

  7. Lima MCP, Simoes S, Pires P, Faneca H, Duzgunes N (2001) Cationic lipid-DNA complexes in gene delivery: from biophysics to biological applications. Adv Drug Deliv Rev 47:277–294

    Article  Google Scholar 

  8. Felgner JH, Kumar R, Sridhar CN, Wheeler CJ, Tsai YJ, Border R et al (1994) Enhanced gene delivery and mechanism studies with a novel series of cationic lipid formulations. J Biol Chem 269:2550–2561

    CAS  PubMed  Google Scholar 

  9. Sternberg B, Hong K, Zheng W, Papahadjopoulos D (1998) Ultrastructural characterization of cationic liposome-DNA complexes showing enhanced stability in serum and high transfection activity in vivo. Biochim Biophys Acta 1375:23–35

    Article  CAS  PubMed  Google Scholar 

  10. Li CX, Parker A, Menocal E, Xiang S, Borodyansky L, Fruehauf JH (2006) Delivery of RNA interference. Cell Cycle 5:2103–2109

    Article  CAS  PubMed  Google Scholar 

  11. Spagnou S, Miller AD, Keller M (2004) Lipidic carriers of siRNA: differences in the formulation, cellular uptake, and delivery with plasmid DNA. Biochemistry 43:13348–13356

    Article  CAS  PubMed  Google Scholar 

  12. Li W, Ishida T, Tachibana R, Almofti MR, Wang X, Kiwada H (2004) Cell type-specific gene expression, mediated by TFL-3, a cationic liposomal vector, is controlled by a post-transcription process of delivered plasmid DNA. Int J Pharm 276:67–74

    Article  CAS  PubMed  Google Scholar 

  13. Almofti MR, Harashima H, Shinohara Y, Almofti A, Li W, Kiwada H (2003) Lipoplex size determines lipofection efficiency with or without serum. Mol Membr Biol 20:35–43

    Article  CAS  PubMed  Google Scholar 

  14. Hirsch-Lerner D, Zhang M, Eliyahu H, Ferrari ME, Wheeler CJ, Barenholz Y (2005) Effect of “helper lipid” on lipoplex electrostatics. Biochim Biophys Acta 1714:71–84

    Article  CAS  PubMed  Google Scholar 

  15. Hofland HEJ, Shephard L, Sullivan SM (1996) Formation of stable cationic lipid/DNA complexes for gene transfer. Proc Natl Acad Sci USA 93:7305–7309

    Article  CAS  PubMed  Google Scholar 

  16. Boussif O, Lezoualc’h F, Zanta MA, Mergny MD, Scherman D, Demeneix B, Behr J-P (1995) A versatile vector for gene and oligonucleotide transfer into cells in culture and in vivo: Polyethylenimide. Proc Natl Acad Sci USA 92:7297–7301

    Article  CAS  PubMed  Google Scholar 

  17. Doyle SR, Chan CK (2007) Differential intracellular distribution of DNA complexed with polyethylenimine (PEI) and PEI-polyarginine PTD influences exogenous gene expression within live COS-7 cells. Genet Vaccines Ther 5:11

    Article  PubMed  Google Scholar 

  18. Lleres D, Weibel JM, Heissler D, Zuber G, Duportail G, Mely Y (2004) Dependence of the cellular internalization and transfection efficiency on the structure and physicochemical properties of cationic detergent/DNA/liposomes. J Gene Med 6:415–428

    Article  CAS  PubMed  Google Scholar 

  19. Zuidam NJ, Hirsch-Lerner D, Margulies S, Barenholz Y (1999) Lamellarity of cationic liposomes and mode of preparation of lipoplexes affect transfection efficiency. Biochim Biophys Acta 1419:207–220

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

The research was supported in part by the Health and Labour Sciences Research Grants for Research on Advanced Medical Technology from The Ministry of Health, Labour and Welfare of Japan. We thank the Japan Association for the Advancement of Medical Equipment for supporting the Postdoctoral Fellowship for Dr. Jose Mario Barichello.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Hiroshi Kiwada .

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2010 Humana Press, a part of Springer Science+Business Media, LLC

About this protocol

Cite this protocol

Barichello, J.M., Ishida, T., Kiwada, H. (2010). Complexation of siRNA and pDNA with Cationic Liposomes: The Important Aspects in Lipoplex Preparation. In: Weissig, V. (eds) Liposomes. Methods in Molecular Biology, vol 605. Humana Press. https://doi.org/10.1007/978-1-60327-360-2_32

Download citation

  • DOI: https://doi.org/10.1007/978-1-60327-360-2_32

  • Published:

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-60327-359-6

  • Online ISBN: 978-1-60327-360-2

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics