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Related Concept Videos

Ribozymes02:47

Ribozymes

The term ribozyme is used for RNA that can act as an enzyme. Ribozymes are mainly found in selected viruses, bacteria, plant organelles, and lower eukaryotes. Ribozymes were first discovered in 1982 when Tom Cech’s laboratory observed Group I introns acting as enzymes. This was shortly followed by the discovery of another ribozyme, Ribonulcease P, by Sid Altman’s laboratory. Both Cech and Altman received the Nobel Prize in chemistry in 1989 for their work on ribozymes.
Ribozymes can be...
Ribozymes02:47

Ribozymes

The term ribozyme is used for RNA that can act as an enzyme. Ribozymes are mainly found in selected viruses, bacteria, plant organelles, and lower eukaryotes. Ribozymes were first discovered in 1982 when Tom Cech’s laboratory observed Group I introns acting as enzymes. This was shortly followed by the discovery of another ribozyme, Ribonulcease P, by Sid Altman’s laboratory. Both Cech and Altman received the Nobel Prize in chemistry in 1989 for their work on ribozymes.
Ribozymes can be...
Ribosome Profiling02:24

Ribosome Profiling

Ribosome profiling or ribo-sequencing is a deep sequencing technique that produces a snapshot of active translation in a cell. It selectively sequences the mRNAs protected by ribosomes to get an insight into a cell’s translation landscape at any given point in time.
Applications of ribosome profiling
Ribosome profiling has many applications, including in vivo monitoring of translation inside a particular organ or tissue type and quantifying new protein synthesis levels.
The technique helps...
Riboswitches01:56

Riboswitches

Riboswitches are non-coding mRNA domains that regulate the transcription and translation of downstream genes without the help of proteins. Riboswitches bind directly to a metabolite and can form unique stem-loop or hairpin structures in response to the amount of the metabolite present. They have two distinct regions – a metabolite-binding aptamer and an expression platform.
The aptamer has high specificity for a particular metabolite which allows riboswitches to specifically regulate...
Experimental RNAi02:15

Experimental RNAi

RNA interference (RNAi) is a cellular mechanism that inhibits gene expression by suppressing its transcription or activating the RNA degradation process. The mechanism was discovered by Andrew Fire and Craig Mello in 1998 in plants. Today, it is observed in almost all eukaryotes, including protozoa, flies, nematodes, insects, parasites, and mammals. This precise cellular mechanism of gene silencing has been developed into a technique that provides an efficient way to identify and determine the...

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Direct selection for ribozyme cleavage activity in cells.

Xi Chen1, Lisa Denison, Matthew Levy

  • 1Department of Chemistry and Biochemistry, University of Texas at Austin, Austin, Texas 78712, USA.

RNA (New York, N.Y.)
|September 25, 2009
PubMed
Summary
This summary is machine-generated.

This study introduces a hybrid in vitro/in vivo selection method for creating functional RNAs. This approach generates novel hammerhead ribozymes that inhibit gene expression in vivo.

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Area of Science:

  • Molecular Biology
  • RNA Therapeutics
  • Genetic Engineering

Background:

  • Traditional RNA selection methods face limitations, including non-physiological conditions in vitro and small pool sizes in vivo.
  • Developing RNA molecules for in vivo genetic regulation requires selection schemes that accurately mimic cellular environments.

Purpose of the Study:

  • To develop a novel hybrid selection strategy that combines the advantages of in vitro and in vivo methods.
  • To generate functional hammerhead ribozymes capable of in vivo gene expression inhibition.

Main Methods:

  • A hybrid selection scheme involving PCR-amplified expression templates transfected into mammalian cells.
  • In vivo transcription, hammerhead RNA self-cleavage, and subsequent amplification of functional ribozyme species.
  • Development of a quantitative kinetic model to evaluate selection stringency.

Main Results:

  • Successful selection of multiple cis-cleaving hammerhead ribozyme variants functional in vivo.
  • Demonstrated inhibition of gene expression by the selected ribozyme variants.
  • Establishment of a kinetic model for optimizing hybrid selection protocols.

Conclusions:

  • The hybrid selection method effectively generates functional hammerhead ribozymes for in vivo applications.
  • Selected ribozymes offer a promising avenue for gene expression regulation.
  • The developed kinetic model aids in refining selection strategies and guiding future research.