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Ribozymes pp 253-267 | Cite as

Cloning and Detection of Aptamer-Ribozyme Conjugations

  • Ryan P. Goguen
  • Anne GatignolEmail author
  • Robert J. ScarboroughEmail author
Protocol
  • 71 Downloads
Part of the Methods in Molecular Biology book series (MIMB, volume 2167)

Abstract

RNA aptamers can be used to target proteins or nucleic acids for therapeutic purposes and are candidates for RNA-mediated gene therapy. Like other small therapeutic RNAs, they can be expressed in cells from DNA templates that include a cellular promoter upstream of the RNA coding sequence. Secondary structures flanking aptamers can be used to enhance the activity or stability of these molecules. Notably, flanking self-cleaving ribozymes to remove extraneous nucleotides included during transcription as well as flanking hairpins to improve RNA stability have been used to increase the effect of therapeutic aptamers. Here we describe the cloning procedure of aptamers containing different flanking secondary structures and methods to compare their expression levels by a northern blot protocol optimized for the detection of small RNA molecules.

Key words

RNA aptamers Aptamer-ribozyme conjugations Northern blot Overlapping PCR Small RNAs Self-cleavage RNA stability 

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2021

Authors and Affiliations

  1. 1.Lady Davis Institute for Medical ResearchMontréalCanada
  2. 2.Department of Microbiology and ImmunologyMcGill UniversityMontréalCanada
  3. 3.Department of Medicine, Division of Experimental MedicineMcGill UniversityMontréalCanada

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