miRNA antagonists (i.e. LNA) have been shown as a very efficient technique to downregulate the expression of microRNA in insects, via hemocele injection. Does anyone knows if the same technic was used with miRNA mimics? If yes, was it done with precursors to miRNA, double strand mimics or single strand mimics?
We have microinjected Antagomir and pre-mir into newborn mice using the in vivo jetPEI solution. We found the injected oligonucleotides to be taken up by surrounding tissues, resulting in significant changes in levels of mRNAs and miRNAs .
Dear Harald,
Is it possible to use the same system, jetPEI, for brain injection???
I think that the open circulatory system of arthropods could be a problem. Each microRNA have a precise level at each tissue. Probably, microinjections in the hemocele modify the concentration of this particular RNA in the whole insect. If you are looking for a precise phenotypic effect, this particula phenotype will be lost by the pleiotropic effect.
Another interesting point in arthropods is the percentage of loading of each particular strand of the microRNA in AGO1 and AGO2 and the different effect that each one have in the mRNA regulation, blocking the translation or cleaving the messenger. We could be sure about the appropriate loading of the strands?
Dear Vincenzo,
This should be possible, although we have not tried. There is a an in vivo transfection medium which specifically appears to be designed for the brain (jetSI® 10 mM.). Visit http://www.polyplus-transfection.com.
Ladys(no one ?) and Gentle men
My fellow is also working on a miRNA injiected topic, but we use zebrafish as research object. I am wondering that is there any specific miRNA antagonists or miRNA mimics can be synthetized to our sample miRNA? Is this technique has been proved precisely credible? How can I get them? The miRNA mimics or miRNA antagonists may help a lot for us to find out which kind of miRNA we injected into fish embryos are the key-fuctional one, or they maybe have a complex regulate-relationship among each others.
I am new in this kind of research area. Any advice is vital for me and thanks a lot for that.
Perhaps antagomirs may be useful for your experiments? Look at the Ambion web-site.
Thank you all for your answers.
Coming back to the mimics topic, does anyone know if the synthesis of the miRNA precursor could be done by doing the RNA strand of the miRNA precursor in our lab? it will fold as a precursor and it will be processed ? Or it's better to buy a comercial one?
Instead of using AMOs (Anti-miRNA Oligonucleotides) can we use some kind of chemicals that directly influence miRNA level and hence causing its suppression /overexpression??
Unlikely in my view. You are here looking for a sequence-specific effect. Your controls will be injections of a corresponding RNA molecule, but with a scrambled sequence.
I have heard some about RNAi and RNAa mechanisms. As you know miRNAs are mostly regulated via epigenetics which can be stable. By referring to the published techniques you can make a candidate miRNA down- or up-regulated.
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