Ribozymes and siRNA protocols

1. Front Matter

   Pages i-xv

   PDF

2. Ribozyme- and siRNA-Mediated mRNA Degradation

   • Mouldy Sioud

   Pages 1-8

3. Combination of Chemical and Enzymatic RNA Synthesis

   • Rajesh K. Gaur, Andreas Hanne, Guido Krupp

   Pages 9-17

4. Determination of Kinetic Parameters for Hammerhead and Hairpin Ribozymes

   • Martha J. Fedor

   Pages 19-32

5. Real-Time Monitoring of RNA and DNA Reactions by Fluorescence Detection

   • Kumud K. Singh, Andreas Hanne, Guido Krupp

   Pages 33-48

6. Quantification of Ribozyme Target RNA Using Real-Time PCR

   • Dagmar Klein, Camillo Ricordi, Ricardo L. Pastori

   Pages 49-56

7. Analysis of Ribozyme Structure and Function by Nucleotide Analog Interference Mapping

   • Soumitra Basu, Catherine Pazsint, Gopeswar Chowdhury

   Pages 57-75

8. Analysis of Global Conformational Transitions in Ribozymes

   • David M. J. Lilley

   Pages 77-108

9. In Vivo Detection of Ribozyme Cleavage Products and RNA Structure by Use of Terminal Transferase-Dependent PCR

   • Hsiu-Hua Chen, Daniela Castanotto, Jeanne LeBon, John J. Rossi, Arthur D. Riggs

   Pages 109-123

10. Identification of Efficient Cleavage Sites in Long-Target RNAs

    • Wei-Hua Pan, Gary A. Clawson

    Pages 125-144

11. Selection In Vitro of Allosteric Ribozymes

    • Adam Roth, Ronald R. Breaker

    Pages 145-164

12. Regulation of Ribozyme Cleavage Activity by Oligonucleotides

    • Yasuo Komatsu, Eiko Ohtsuka

    Pages 165-177

13. Tetracycline-Regulated Expression of Hammerhead Ribozymes In Vivo

    • Emma T. Bowden, Anna T. Riegel

    Pages 179-194

14. Ribozyme Expression Systems

    • Masayuki Sano, Kazunari Taira

    Pages 195-207

15. Design and Expression of Chimeric U1/Ribozyme Transgenes

    • Roger Abounader, Robert Montgomery, Harry Dietz, John Laterra

    Pages 209-219

16. Design and Validation of Therapeutic Hammerhead Ribozymes for Autosomal Dominant Diseases

    • Jason J. Fritz, Marina Gorbatyuk, Alfred S. Lewin, William W. Hauswirth

    Pages 221-236

17. Helicase-Attached Novel Hybrid Ribozymes

    • Hiroaki Kawasaki, Masaki Warashina, Tomoko Kuwabara, Kazunari Taira

    Pages 237-243

18. Functional Gene Discovery Using Hybrid Ribozyme Libraries

    • Yoshio Kato, Masaru Tsunemi, Makoto Miyagishi, Hiroaki Kawasaki, Kazunari Taira

    Pages 245-256

19. Maxizyme Technology

    • Mayu Iyo, Hiroaki Kawasaki, Kazunari Taira

    Pages 257-265

20. Target-Site Selection for the 10–23 DNAzyme

    • Murray J. Cairns, Lun-Quan Sun

    Pages 267-277

In this completely updated and expanded edition of a classic bench manual, hands-on experts take advantage of the latest advances in ribozyme, DNAzyme, hammerhead ribozymes and derivatives, and RNA interference technologies to describe in detail the exciting and successful methods now available for gene inactivation in vitro and in vivo. Their optimized techniques employ hairpin ribozymes, DNAzymes, hammerhead ribozymes and derivatives, group I intron ribozymes, RNase P ribozymes, and siRNAs, as well as general methods for RNA structure analysis, delivery of oligonucleotides, and gene therapy. Also provided are novel methods for identifying accessible cellular mRNA sites; group I intron and RNAse P ribozyme protocols for effective design, selection, and therapeutic applications; and the latest RNAi methods for sequence-specific gene silencing in a wide variety of organisms. Additional techniques cover the analysis of ribozyme structures and conformational transitions using nucleotide analog interference mapping and fluorescence resonance energy transfer, the use of ribozymes in clinical and gene therapy, and the use of ribozymes and DNAzymes in rodent models of human disease. Each proven protocol includes a background introduction outlining the principle behind the technique, step-by-step instructions, lists of equipment and reagents, and tips on troubleshooting and avoiding known pitfalls.
Comprehensive and up-to-date, Ribozymes and siRNA Protocols details for experienced and novice investigators alike the many exciting advances in our understanding of nucleic acid enzymes, as well as demonstrating how they may be used to analyze gene function and target validation, and to productively develop novel therapeutics for human diseases.

"...a timely and important addition to the literature in this rapidly expanding field...The vast scope of the book will appeal to experienced researchers as a source for new avenues of study, and as a central reference for the latest technologies."
-Biochemist

"...will be very useful for both novices and experts as a comprehensive collection of ribozyme protocols..." -Doody's Health Sciences Book Review Journal

"...will provide a valuable source of reference for many laboratories..."
-Microbiology Today

• Institute of Cancer Research, The Norwegian Radium Hospital, Oslo, Norway

  Mouldy Sioud

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