siRNA in vivo use - (Jul/23/2006 )

Hi all
I have a question about siRNA transfection in vivo. We are currently trying to get successful siRNA transfection into ganglion cells. As far as I am aware there has been nothing published on delivery into this tissue. We are using the Invitrogen range of products including BlockIt siRNA-FITC and Lipofectamine 2000. We have tried a few injections of varying concentrations (and times) of the BlockIt with no transfection. We can see fluorescence in the ganglion, but no transfection into the cell bodies.
Has anyone used the Invitrogen products in vivo, or had success with siRNA in vivo injections that could give me a few pointers.

Any suggestions appreciated!

-nicolejodie-

I dont know about siRNA delivery to nerve system, but PEI has been used to deliver genes to CNS. See : Mol Ther. 2001 May;3(5 Pt 1):658-64.

AAV can also do very good gene delivery to CNS.

Maybe you can think about shRNA/ viral or non-viral strategy?

-genehunter-1-

I have used AAV for shRNA invivo (CNS) and works very good. I am starting the same with Lenti and it works good so far on cell culture. I have to try it invivo. But people have been using it for invivo (CNS0 and is also good.

-scolix-

I hate to seem really dumb, but as I'm new to the siRNA scene I have to ask what does AAV stand for? I'm still getting grips on the terminology used in this field.
Thanks

-nicolejodie-

AAV = adeno associated virus

-genehunter-1-

Thanks for that!

Eventually we will be moving to viral injection, but while we are waiting for approval, thought it might be nice to give in vivo siRNA transfection a go.
So, if anyone has any other suggestions for me that would be great!

-nicolejodie-

One of my colleagues used to inject siRNA into retinal ganglion cells (intravitreal space). the transfection isnt great but it depends on what sort of experiments one has planned.

-scolix-

We want to do in utero electroporation with shRNA into the mouse brain, but which vector would be the best to use? We were thinking of the pSUPER or the pSilencer , but all suggestions are welcome.

-lautje-

QUOTE (lautje @ Nov 1 2006, 02:56 PM)

We want to do in utero electroporation with shRNA into the mouse brain, but which vector would be the best to use? We were thinking of the pSUPER or the pSilencer , but all suggestions are welcome.

Hello,

Did you get any success with the in utero EP? I am just starting to get into this but find that I never come even close to the 80% efficiency all the papers talk about and I am using a clontech pCMV-eGFP for the test runs so that should not be the problem.
What concentration of the plasmid did you use with best results?

Best

chili

-chili-