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A hammerhead ribozyme substrate and reporter for in vitro kinetoplastid RNA editing

Published online by Cambridge University Press:  24 April 2002

BINGBING WANG ,
REZA SALAVATI ,
STEFAN HEIDMANN  and
KENNETH STUART
BINGBING WANG
Affiliation:
Department of Pathobiology, University of Washington, Seattle, Washington 98195, USA Seattle Biomedical Research Institute, Seattle, Washington 98109, USA
REZA SALAVATI
Affiliation:
Department of Pathobiology, University of Washington, Seattle, Washington 98195, USA Seattle Biomedical Research Institute, Seattle, Washington 98109, USA
STEFAN HEIDMANN
Affiliation:
Department of Genetics, University of Bayreuth, 95440 Bayreuth, Germany
KENNETH STUART
Affiliation:
Department of Pathobiology, University of Washington, Seattle, Washington 98195, USA Seattle Biomedical Research Institute, Seattle, Washington 98109, USA
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Abstract

Current in vitro assays for RNA editing in kinetoplastids directly examine the products generated by incubation of pre-mRNA substrate with guide RNA (gRNA) and mitochondrial (mt) extract. RNA editing substrates that are modeled on hammerhead ribozymes were designed with catalytic cores that contained or lacked additional uridylates (Us). They proved to be sensitive reporters of editing activity when used for in vitro assays. A deletion editing substrate that is based on A6 pre-mRNA had no ribozyme activity, but its incubation with gRNA and mt extract resulted in its deletion editing and production of a catalytically active ribozyme. Hammerhead ribozymes are thus sensitive tools to assay in vitro RNA editing.

Keywords

gRNATrypanosoma bruceiuridylate deletionuridylate insertion
Type
METHODS
Information
RNA , Volume 8 , Issue 4 , April 2002 , pp. 548 - 554
Copyright
2002 RNA Society

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