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Viral delivery of small-hairpin RNAs for reducing gene expression in the rodent brain.

Lasek AW, Heberlein U - Alcohol Res Health (2008)

Bottom Line: This article reviews a strategy to ensure long-term expression of a type of interfering RNA molecule-small-hairpin RNAs (shRNAs)-through the use of viral delivery systems (i.e., vectors).Expression of these shRNAs leads to the destruction of the intermediary molecules (i.e., messenger RNA [mRNA] molecules) generated during the expression of the target gene.

View Article: PubMed Central - PubMed

Affiliation: Ernest Gallo Clinic and Research Center at the University of California at San Francisco, Emeryville, California.

ABSTRACT
This article reviews a strategy to ensure long-term expression of a type of interfering RNA molecule-small-hairpin RNAs (shRNAs)-through the use of viral delivery systems (i.e., vectors). Expression of these shRNAs leads to the destruction of the intermediary molecules (i.e., messenger RNA [mRNA] molecules) generated during the expression of the target gene.

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Related in: MedlinePlus

Example of a neuron located in the mouse nucleus accumbens that has been infected by a lentivirus carrying a small-hairpin RNA (shRNA). The success of the infection is demonstrated by the green fluorescence of the cell, which stems from a marker gene (i.e., green fluorescent protein [GFP]) that was introduced into the cell via the same virus as the shRNA. The image was taken 2 weeks after the infection. The inset shows a higher magnification view of the dashed rectangle, illustrating GFP expression even in the tiny extensions protruding from the neuron.
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f13-arh-31-3-259: Example of a neuron located in the mouse nucleus accumbens that has been infected by a lentivirus carrying a small-hairpin RNA (shRNA). The success of the infection is demonstrated by the green fluorescence of the cell, which stems from a marker gene (i.e., green fluorescent protein [GFP]) that was introduced into the cell via the same virus as the shRNA. The image was taken 2 weeks after the infection. The inset shows a higher magnification view of the dashed rectangle, illustrating GFP expression even in the tiny extensions protruding from the neuron.

Mentions: After efficient shRNA sequences have been identified, the corresponding modified virus is prepared in larger quantities (e.g., using commercially available kits) and purified. It then is injected into the brain region of interest, and knock-down of the target gene can be assessed approximately 2 weeks later (see figure 13). Initial analyses then serve to determine the time frame of the gene knock-down (i.e., when exactly and for how long knock-down is strongest). Once the timeframe of gene knock-down in the living animal (i.e., in vivo) has been determined, additional animals are infected with the modified lentivirus and their behavior tested using appropriate tests. When these behavioral analyses are completed, the site and extent of infection are determined by staining the appropriate brain sections or treating them with immune molecules (i.e., antibodies) that make cells expressing the GFP marker gene visible (see figure 14).


Viral delivery of small-hairpin RNAs for reducing gene expression in the rodent brain.

Lasek AW, Heberlein U - Alcohol Res Health (2008)

Example of a neuron located in the mouse nucleus accumbens that has been infected by a lentivirus carrying a small-hairpin RNA (shRNA). The success of the infection is demonstrated by the green fluorescence of the cell, which stems from a marker gene (i.e., green fluorescent protein [GFP]) that was introduced into the cell via the same virus as the shRNA. The image was taken 2 weeks after the infection. The inset shows a higher magnification view of the dashed rectangle, illustrating GFP expression even in the tiny extensions protruding from the neuron.
© Copyright Policy - public-domain
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3860491&req=5

f13-arh-31-3-259: Example of a neuron located in the mouse nucleus accumbens that has been infected by a lentivirus carrying a small-hairpin RNA (shRNA). The success of the infection is demonstrated by the green fluorescence of the cell, which stems from a marker gene (i.e., green fluorescent protein [GFP]) that was introduced into the cell via the same virus as the shRNA. The image was taken 2 weeks after the infection. The inset shows a higher magnification view of the dashed rectangle, illustrating GFP expression even in the tiny extensions protruding from the neuron.
Mentions: After efficient shRNA sequences have been identified, the corresponding modified virus is prepared in larger quantities (e.g., using commercially available kits) and purified. It then is injected into the brain region of interest, and knock-down of the target gene can be assessed approximately 2 weeks later (see figure 13). Initial analyses then serve to determine the time frame of the gene knock-down (i.e., when exactly and for how long knock-down is strongest). Once the timeframe of gene knock-down in the living animal (i.e., in vivo) has been determined, additional animals are infected with the modified lentivirus and their behavior tested using appropriate tests. When these behavioral analyses are completed, the site and extent of infection are determined by staining the appropriate brain sections or treating them with immune molecules (i.e., antibodies) that make cells expressing the GFP marker gene visible (see figure 14).

Bottom Line: This article reviews a strategy to ensure long-term expression of a type of interfering RNA molecule-small-hairpin RNAs (shRNAs)-through the use of viral delivery systems (i.e., vectors).Expression of these shRNAs leads to the destruction of the intermediary molecules (i.e., messenger RNA [mRNA] molecules) generated during the expression of the target gene.

View Article: PubMed Central - PubMed

Affiliation: Ernest Gallo Clinic and Research Center at the University of California at San Francisco, Emeryville, California.

ABSTRACT
This article reviews a strategy to ensure long-term expression of a type of interfering RNA molecule-small-hairpin RNAs (shRNAs)-through the use of viral delivery systems (i.e., vectors). Expression of these shRNAs leads to the destruction of the intermediary molecules (i.e., messenger RNA [mRNA] molecules) generated during the expression of the target gene.

Show MeSH
Related in: MedlinePlus