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Transfection - Plasmid Uptake Control


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#1 Oxford

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Posted 06 January 2010 - 08:26 AM

Hi,

I'm transfecting HEK 293T cells by nucleofection and I want a control for the amount of plasmid taken up in each transfection.

I've read that this can be done by Southern blot but isn't very quantitative and is an out-dated technique.

The alternatives I've come across are cotransfections with reporter plasmids, but surely plasmid-reporter plasmid interactions and different capactity for uptake between reporter and construct could skew results?

I was wondering what is generally done in the field?

Thanks for your time,

Adam

#2 madrius1

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Posted 06 January 2010 - 09:45 AM

What about quantitative PCR with a standard curve?

#3 laurequillo

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Posted 07 January 2010 - 01:38 AM

You mean to check the efficiency of the electroporation, right? ( I dont know if I got it right).
You can use a plasmid producing gfp and just check the amount of green cells (IF or cytometry)
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