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Ribozymes pp 21-40 | Cite as

Mechanistic Analysis of the Hepatitis Delta Virus (HDV) Ribozyme: Methods for RNA Preparation, Structure Mapping, Solvent Isotope Effects, and Co-transcriptional Cleavage

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Part of the Methods in Molecular Biology book series (MIMB, volume 848)

Abstract

Small ribozymes such as the hairpin, hammerhead, VS, glm S, and hepatitis delta virus (HDV) are self-cleaving RNAs that are typically characterized by kinetics and structural methods. Working with these RNAs requires attention to numerous experimental details. In this chapter we focus on four different experimental aspects of ribozyme studies: preparing the RNA, mapping its structure with reverse transcription and end-labeled techniques, solvent isotope experiments, and co-transcriptional cleavage assays. Although the focus of these methods is the HDV ribozyme, the methods should be applicable to other ribozymes.

Key words

Co-transcriptional RNA folding pKa Proton inventory Sequencing Solvent isotope 
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Notes

Acknowledgments

We thank NSF MCB-0527102 for supporting this research.

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Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  1. 1.Department of ChemistryThe Pennsylvania State UniversityUniversity ParkUSA
  2. 2.Institute for Bioscience and Biotechnology Research, NISTRockvilleUSA
  3. 3.Department of ChemistryUniversity of North CarolinaChapel HillUSA

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