Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Paper
  • Published:

Minicircle: an improved DNA molecule for in vitro and in vivo gene transfer

Gene Therapy volume 6, pages 209–218 (1999)Cite this article

Abstract

Minicircles are a new form of supercoiled DNA molecule for nonviral gene transfer which have neither bacterial origin of replication nor antibiotic resistance marker. They are thus smaller and potentially safer than the standard plasmids currently used in gene therapy. They were obtained in E. coli by att site-specific recombination mediated by the phage λ integrase, which was used to excise the expression cassette from the unwanted plasmid sequences. We produced two minicircles containing the luciferase or β-galactosidase gene under the control of the strong human cytomegalovirus immediate–early enhancer/promoter. Comparing maximal differences, these minicircles gave 2.5 to 5.5 times more reporter gene activity than the unrecombined plasmid in the NIH3T3 cell line and rabbit smooth muscle cells. Moreover, injection in vivo into mouse cranial tibial muscle, or human head and neck carcinoma grafted in nude mice resulted in 13 to 50 times more reporter gene expression with minicircles than with the unrecombined plasmid or larger plasmids. Histological analysis in muscle showed there were more transfected myofibers with minicircles than with unrecombined plasmid.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7

Similar content being viewed by others

References

  1. Darquet AM et al. A new DNA vehicle for nonviral gene delivery: supercoiled minicircle Gene Therapy 1997 4: 1341–1349

    Article  CAS  PubMed  Google Scholar 

  2. Pang A . Production of antibodies against Bacillus thuringiensis delta-endotoxin by injecting its plasmids Biochem Biophys Res Commun 1994 202: 1227–1234

    Article  CAS  PubMed  Google Scholar 

  3. Valera A, Perales J, Hatzoglou M, Bosch F . Expression of the neomycin-resistance (neo) gene induces alterations in gene expression and metabolism Hum Gene Ther 1994 5: 449–456

    Article  CAS  PubMed  Google Scholar 

  4. Klinman D, Yamshchikov G, Ishigatsubo Y . Contribution of CpG motifs to the immunogenicity of DNA vaccines J Immunol 1997 158: 3635–3639

    CAS  PubMed  Google Scholar 

  5. Pisetsky D . The immunologic properties of DNA J Immunol 1994 156: 421–423

    Google Scholar 

  6. Sato Y et al. Immunostimulatory DNA sequences necessary for effective intradermal gene immunization Science 1996 273: 352–354

    Article  CAS  PubMed  Google Scholar 

  7. Landy A . Dynamic, structural, and regulatory aspects of lambda site-specific recombination Annu Rev Biochem 1989 58: 913–949

    Article  CAS  PubMed  Google Scholar 

  8. Hasan N, Szybalski W . Control of cloned gene expression by promoter inversion in vivo: construction of improved vectors with a multiple cloning site and the Ptac promoter Gene 1987 56: 145–151

    Article  CAS  PubMed  Google Scholar 

  9. Gellert M, Nash H . Communication between segments of DNA during site-specific recombination Nature 1987 325: 401–404

    Article  CAS  PubMed  Google Scholar 

  10. Davis H et al. Plasmid DNA is superior to viral vectors for direct gene transfer into adult mouse skeletal muscle Hum Gene Ther 1993 4: 733–740

    Article  CAS  PubMed  Google Scholar 

  11. Buttrick P, Kass A, Kitsis R . Behavior of genes directly injected into the rat heart in vivo Circ Res 1992 70: 193–198

    Article  CAS  PubMed  Google Scholar 

  12. Ledley F . Non viral gene therapy: the promise of genes as pharmaceutical products Hum Gene Ther 1995 6: 1129–1144

    Article  CAS  PubMed  Google Scholar 

  13. Jain R . Transport of molecules in the tumor interstitium: a review Cancer Res 1987 47: 3039–3051

    CAS  PubMed  Google Scholar 

  14. Levy M, Barron L, Meyer K, Szoka FJ . Characterization of plasmid DNA transfer into mouse skeletal muscle: evaluation of uptake mechanism, expression and secretion of gene products into blood Gene Therapy 1996 3: 201–211

    CAS  PubMed  Google Scholar 

  15. Wolff J et al. Expression of naked plasmids by cultured myotubes and entry of plasmids into T tubules and caveolae of mammalian skeletal muscle J Cell Sci 1992 103: 1249–1259

    CAS  PubMed  Google Scholar 

  16. Friend D, Papahadjopoulos D, Debs R . Endocytosis and intracellular processing accompanying transfection mediated by cationic liposomes Biochim Biophys Acta 1996 1278: 41–50

    Article  PubMed  Google Scholar 

  17. Zabner J et al. Cellular and molecular barriers to gene transfer by a cationic lipid J Biol Chem 1995 270: 18997–19007

    Article  CAS  PubMed  Google Scholar 

  18. Gao X, Huang L . Cationic liposome-mediated gene transfer Gene Therapy 1995 2: 710–722

    CAS  PubMed  Google Scholar 

  19. Danko I et al. Pharmacological enhancement of in vivo foreign gene expression in muscle Gene Therapy 1994 1: 114–121

    CAS  PubMed  Google Scholar 

  20. Dworetzky S, Lanford R, Feldherr C . The effects of variations in the number and sequence of targeting signals on nuclear uptake J Cell Biol 1988 107: 1279–1287

    Article  CAS  PubMed  Google Scholar 

  21. Lanford R, Kanda P, Kennedy R . Induction of nuclear transport with a synthetic peptide homologous to the SV40 T antigen transport signal Cell 1986 46: 575–582

    Article  CAS  PubMed  Google Scholar 

  22. Wils P et al. Efficient purification of plasmid DNA for gene transfer using triple-helix affinity chromatography Gene Therapy 1997 4: 323–330

    Article  CAS  PubMed  Google Scholar 

  23. Sambrook J, Fritsh E, Maniatis T . Molecular Cloning: a Laboratory Manual Cold Spring Harbor Laboratory Press: New York 1989

    Google Scholar 

  24. Sanger F et al. Nucleotide sequence of bacteriohage λ DNA J Mol Biol 1982 162: 729–773

    Article  CAS  PubMed  Google Scholar 

  25. Yanisch-Perron C, Vieira J, Messing J . Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors Gene 1985 33: 103–119

    Article  CAS  PubMed  Google Scholar 

  26. Tanaka M, Herr W . Differential transcriptional activation by Oct-1 and Oct-2: interdependent activation domains induce Oct-2 phosphorylation Cell 1990 60: 375–386

    Article  CAS  PubMed  Google Scholar 

  27. Manthorpe M et al. Gene therapy by intramuscular injection of plasmid ADN: studies on firefly luciferase gene expression in mice Hum Gene Ther 1993 4: 419–431

    Article  CAS  PubMed  Google Scholar 

  28. Sadler J, Tecklenburg M, Betz J . Plasmids containing many tandem copies of a synthetic lactose operator Gene 1980 8: 279–300

    Article  CAS  PubMed  Google Scholar 

  29. Chamley J, Campbell G, McConnell J, Groschel-Stewart U . Comparison of vascular smooth muscle cells from adult human, monkey and rabbit in primary culture and in subculture Cell Tissue Res 1977 177: 503–522

    CAS  PubMed  Google Scholar 

  30. Byk G et al. Synthesis, activity, and structure-activity relationship studies of novel cationic lipids for DNA transfer J Med Chem 1998 41: 224–235

    Article  Google Scholar 

  31. Escriou V et al. Cationic lipid-mediated gene transfer: effectof serum on cellular uptake and intracellular fate of lipopolyamine/DNA complexes Biochim Biophys Acta 1998 1368: 276–288

    Article  CAS  PubMed  Google Scholar 

  32. Wiechelman K, Braun R, Fitzpatrick J . Investigation of the bicinchoninic acid protein assay: identification of the groups responsible for color formation Anal Biochem 1988 175: 231–237

    Article  CAS  PubMed  Google Scholar 

  33. Wolff J et al. Conditions affecting direct gene transfer into rodent muscle in vivo Biotechniques 1991 11: 474–485

    CAS  PubMed  Google Scholar 

  34. Cohen-Tannoudji M, Morello D, Babinet C . Unexpected position-dependent expression of H-2 and beta 2-micro-globulin/lacZ transgenes Mol Reprod Dev 1992 33: 149–159

    Article  CAS  PubMed  Google Scholar 

  35. Davis H, Whalen R, Demeneix B . Direct gene transfer into skeletal muscle in vivo: factors affecting efficiency of transfer and stability of expression Hum Gene Ther 1993 4: 151–159

    Article  PubMed  Google Scholar 

  36. Tissenbaum HA, Parry DJ . The effect of partail denervation of tibialis anterior (TA) muscle on the number and size of motorneurons in TA motornucleus of normal and dystrophic (C57BL dy2j/dy2j) mice Can J Physiol Pharmacol 1991 69: 1769–1773

    Article  CAS  PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. UMR 133 CNRS/Rhône-Poulenc Rorer, Centre de Recherche de Vitry-Alfortville, Vitry sur Seine, France

    A-M Darquet, R Rangara, P Kreiss, B Schwartz & D Scherman

  2. Rhône-Poulenc Rorer Gencell, Centre de Recherche de Vitry-Alfortville, Vitry sur Seine, France

    S Naimi, P Delaère & J Crouzet

Authors
  1. A-M Darquet
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. R Rangara
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. P Kreiss
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. B Schwartz
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. S Naimi
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. P Delaère
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. J Crouzet
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. D Scherman
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Darquet, AM., Rangara, R., Kreiss, P. et al. Minicircle: an improved DNA molecule for in vitro and in vivo gene transfer. Gene Ther 6, 209–218 (1999). https://doi.org/10.1038/sj.gt.3300816

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/sj.gt.3300816

Keywords

This article is cited by

Search

Advanced search

Quick links