Skip to main content

A PCR-Based Approach to Hammerhead Ribozyme Synthesis

  • Protocol
Book cover Therapeutic Applications of Ribozymes

Part of the book series: Methods in Molecular Medicine™ ((MIMM,volume 11))

  • 187 Accesses

Abstract

Inactivation of gene expression by antisense mechanisms in general and by ribozymes in particular show great promise as a research tool as well as therapeutic agent in selectively reducing mRNA species and thus decreasing subsequent protein synthesis (for reviews see refs. 1 and 2).

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 84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 109.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 109.99
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. Marschall, P., Thomson, J B., and Eckstein, F (1994) Inhibition of gene expression with ribozymes. Cell. Mol NeuroBiol 14, 523–538.

    Article  PubMed  CAS  Google Scholar 

  2. Rossi, J. J. (1995) Controlled, targeted, intracellular expression of ribozymes progress and problems. TIBTECH 13, 301–306

    CAS  Google Scholar 

  3. Cameron, F H. and Jennings, P. A. (1989) Specific gene suppression by engineered ribozymes in monkey cells. Proc Natl. Acad Sci USA 86, 9139–9143

    Article  PubMed  CAS  Google Scholar 

  4. Sarver, N, Cantin, E. M., Chang, P S., Zaia, J. A., Ladne, P A, Stephens, D A, and Rossi, J J. (1990) Ribozymes as potential anti-HIV-1 therapeutic agents Science 247, 1222–1225.

    Article  PubMed  CAS  Google Scholar 

  5. Ohta, Y., Kijima, H., Ohkawa, T., Kashini-Sabeth, M., and Scanlon, K J (1996) Tissue-specific expression of an anti-ras ribozyme inhibits proliferation of human malignant melanoma cells. Nucleic Acids Res 24, 938–942.

    Article  PubMed  CAS  Google Scholar 

  6. Sioud, M., Natvig, J. B., and Forre, O (1992) Preformed ribozyme destroys tumour necrosis factor mRNA in human cells J, Mol Biol 223, 831–835.

    Article  CAS  Google Scholar 

  7. Kiehntopf, M, Brach, M A, Licht, T, Petschauer, S, Karawajew, L, Kirschning, C, and Herrmann, F. (1994) Ribozyme-mediated cleavage of the MDR-1 transcript restores chemosensitivity in previously resistant cancer cells. EMBO J 13, 4645–4652.

    PubMed  CAS  Google Scholar 

  8. Leopold, L H., Shore, S K, Newkirk, T 0, Reddy, R M. V, and Reddy, E. P. (1995) Multi-unit ribozyme-mediated cleavage of bcr-abl mRNA in myeloid leukemias. Blood 85, 2162–2170.

    PubMed  CAS  Google Scholar 

  9. Pieken, W A., Olsen, D B, Benseler, F, Aurup, H., and Eckstein, F. (1991) Kinetic characterization of ribonuclease-resistant 2′-modified hammerhead ribozymes. Science 253, 314–317.

    Article  PubMed  CAS  Google Scholar 

  10. Hoffman, L M and Johnson, M G (1994) Enzymatic synthesis of milligram quantities of ribozymes in small volumes. BioTechniques 17, 372–375

    PubMed  CAS  Google Scholar 

  11. Uhlenbeck, O C (1987) A small catalytic oligoribonucleotide. Nature 328, 596–600

    Article  PubMed  CAS  Google Scholar 

  12. Fedor, M J and Uhlenbeck, O C. (1990) Substrate sequence effect on “hammerhead” RNA catalytic efficiency. Proc Natl Acad Sci USA 87, 1668–1672

    Article  PubMed  CAS  Google Scholar 

  13. Milligan, J F, Groebe, D. R, Witherell, G W, and Uhlenbeck, O. C (1987) Oligoribonucleotide synthesis using T7 RNA polymerase and synthetic DNA templates Nucleic Acids Res 15, 8783–8798

    Article  PubMed  CAS  Google Scholar 

  14. Scanlon, K J, Jiao, L., Funato, T, Wang, W., Tone, T, Rossi, J. J., and KashaniSabet, M (1991) Ribozyme-mediated cleavage of c-fos mRNA reduces gene expression of DNA synthesis enzymes and metallothionein. Proc. Natl Acad Sci USA 88, 10,591–10,595

    Article  PubMed  CAS  Google Scholar 

  15. Sane, D C and Deidra, J C (1995) PCR-based production of a ribozyme to plasminogen activator inhibitor-1. Bio Techniques 18, 208–210

    CAS  Google Scholar 

  16. Haseloff, J and Gerlach, W. L. (1988) Simple RNA enzymes with new and highly specific endoribonuclease activities Nature 334, 585–591

    Article  PubMed  CAS  Google Scholar 

  17. Roldan, A. L, Cubellis, M V., Masucci, M. T., Behrendt, N., Lund, L R, Dano, K, Appella, E., and Blasi, F. (1990) Cloning and expression of the receptor for human urokinase plasminogen activator, a central molecule in cell surface plasmin dependent proteolysis. EMBO J 9, 467–474

    PubMed  CAS  Google Scholar 

  18. Ausubel, F M, Brent, R, Kingston, R E, Moore, D D., Seidmar, J G, Smith, J A., Struhl, K, eds (1992) Current Protocols in Molecular Biology. New York Wiley Interscience.

    Google Scholar 

  19. Czerny, T. (1996) High primer concentration improves PCR amplification from random pools. Nucleic Acids Res 24, 985–986

    Article  PubMed  CAS  Google Scholar 

  20. Liang, P and Pardee, A B (1992) Differential display of eukariotic messenger RNA by means of the polymerase chain reaction. Science 257, 967–971.

    Article  PubMed  CAS  Google Scholar 

  21. Lyamichev, V., Brow, M.A.D, and Dahlberg, J E (1993) Structure-specific endonucleolytic cleavage of nucleic acids by eubacterial DNA polymerases. Science 260, 778–783

    Article  PubMed  CAS  Google Scholar 

  22. Karikó, K., Megyeri, K, Xiao, Q, and Barnathan, E B (1994) Lipofectin-aided cell delivery of ribozyme targeted to human urokinase receptor mRNA. FEBS Lett 352, 41–44.

    Article  PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 1998 Humana Press Inc.

About this protocol

Cite this protocol

Karikó, K., Barnathan, E.S. (1998). A PCR-Based Approach to Hammerhead Ribozyme Synthesis. In: Scanlon, K.J. (eds) Therapeutic Applications of Ribozymes. Methods in Molecular Medicine™, vol 11. Springer, Totowa, NJ. https://doi.org/10.1385/0-89603-477-1:427

Download citation

  • DOI: https://doi.org/10.1385/0-89603-477-1:427

  • Publisher Name: Springer, Totowa, NJ

  • Print ISBN: 978-0-89603-477-8

  • Online ISBN: 978-1-59259-595-2

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics