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Related Concept Videos

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...
RNA Interference01:23

RNA Interference

RNA interference (RNAi) is a process in which a small non-coding RNA molecule blocks the post-transcriptional expression of a gene by binding to its messenger RNA (mRNA) and preventing the protein from being translated.
This process occurs naturally in cells, often through the activity of genomically-encoded microRNAs. Researchers can take advantage of this mechanism by introducing synthetic RNAs to deactivate specific genes for research or therapeutic purposes. For example, RNAi could be used...
RNA Interference01:23

RNA Interference

RNA interference (RNAi) is a process in which a small non-coding RNA molecule blocks the post-transcriptional expression of a gene by binding to its messenger RNA (mRNA) and preventing the protein from being translated.
This process occurs naturally in cells, often through the activity of genomically-encoded microRNAs. Researchers can take advantage of this mechanism by introducing synthetic RNAs to deactivate specific genes for research or therapeutic purposes. For example, RNAi could be used...
siRNA - Small Interfering RNAs02:30

siRNA - Small Interfering RNAs

Small interfering RNAs, or siRNAs, are short regulatory RNA molecules that can silence genes post-transcriptionally, as well as the transcriptional level in some cases. siRNAs are important for protecting cells against viral infections and silencing transposable genetic elements.
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Small interfering RNAs (siRNA)02:30

Small interfering RNAs (siRNA)

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MicroRNAs

MicroRNA (miRNA) are short, regulatory RNA transcribed from introns (non-coding regions of a gene) or intergenic regions (stretches of DNA present between genes). Several processing steps are required to form biologically active, mature miRNA. The initial transcript, called primary miRNA (pri-mRNA), base-pairs with itself, forming a stem-loop structure. Within the nucleus, an endonuclease enzyme, called Drosha, shortens the stem-loop structure into hairpin-shaped pre-miRNA. After the pre-miRNA...

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MISSION esiRNA for RNAi Screening in Mammalian Cells
15:31

MISSION esiRNA for RNAi Screening in Mammalian Cells

Published on: May 12, 2010

Minimizing off-target effects by using diced siRNAs for RNA interference.

Jason W Myers1, Jen-Tsan Chi, Delquin Gong

  • 1Stanford University School of Medicine, Department of Molecular Pharmacology, 269 West Campus Drive, CCSR Room 3155, Stanford CA 94305-5174, USA.

Journal of Rnai and Gene Silencing : an International Journal of RNA and Gene Targeting Research
|September 23, 2009
PubMed
Summary

Diced small interfering RNA (siRNA) pools exhibit high specificity in gene silencing, unlike synthetic siRNAs which cause significant off-target effects. This complexity diminishes off-target concerns by reducing individual siRNA concentration.

Keywords:
RNAid-siRNAe-siRNAgene silencingmicroarraymitogen activated protein kinase (MAPK)siRNA

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RNA Interference in Aquatic Beetles as a Powerful Tool for Manipulating Gene Expression at Specific Developmental Time Points
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Area of Science:

  • Molecular Biology
  • Genetics
  • Biochemistry

Background:

  • Synthetic small interfering RNAs (siRNAs) can cause substantial off-target effects, impacting gene silencing experiments.
  • Diced siRNA pools, generated from dsRNA, offer an alternative approach for gene silencing.

Purpose of the Study:

  • To compare the off-target effects of diced siRNA pools versus chemically synthesized siRNAs.
  • To investigate whether the complexity of diced siRNA pools exacerbates or mitigates off-target effects.

Main Methods:

  • Chemically synthesized siRNAs and enzymatically diced siRNA pools targeting p38alpha MAPK (MAPK14) were generated.
  • Microarray analysis was used to assess mRNA abundance changes, indicating off-target effects.
  • Synthetic siRNAs were diluted within diced pools to evaluate concentration-dependent effects.

Main Results:

  • Chemically synthesized siRNAs induced off-target changes in hundreds of mRNAs.
  • Diced siRNA pools targeting p38alpha MAPK showed minimal off-target effects.
  • Diluting synthetic siRNAs in diced pools significantly reduced or eliminated off-target effects.

Conclusions:

  • The high complexity of diced siRNA pools reduces the concentration of individual siRNAs, thereby minimizing off-target effects.
  • This finding supports the specificity of RNA interference in organisms using endogenous diced siRNAs.
  • Diced siRNA pools present a strategy to enhance the specificity of RNA interference in mammalian cells.