labeling miRNA - (Mar/26/2008 )

Is it possible to label miRNA with a fluorochrome to see if your transfection in cells with the miRNA was succesfull?

There is a Cy3-labeled negative control Pre-miR from Ambion that I've used for this purpose. It works quite well, can easily see red fluorescence.

The Pre-miRs are synthetic microRNAs (although Ambion won't tell us the structure, they're not actually hairpins, I think they're just chemically modified siRNAs that have the same sequence as the miRNA duplex). The negative control is a non-targeting Pre-miR. It is also available conjugated to Cy3 so that you can check transfection efficiency.

ETA: There are also commercial miRNA labeling kits so that you can label small RNAs with Cy3/Cy5 for microarray studies. I guess you could label your isolated miRNAs with this kit and transfect?

Does there exist a normal PCR where I can form miRNAs with, who are labeled with a fluorochrome?

see
http://www.mirusbio.com/products/labelit/labeling/

but it would cheaper to buy one that is already labeled as miRNA man recommended. The result is same. Transfection reagent does not recognize sequence difference. its ok to use it.

I've tried the simplest method......i made synthetic mature miRNAs labelled with alexa fluor 488.
i just ordered them like normal PCR primers at Invitrogen as RNA primers with U instead of T.
I don't know yet if they work as real mature miRNAs but i can detect them by real time PCR.
It's only a bit more expensive than normal DNA primers but less than commercial labelling kits.
Hope it helps!
Fizban