Multiple plasmid FuGENE 6 transfections

Ian A. York iayork at panix.com
Sat Sep 22 08:48:01 EST 2001


In article <3BACF420.16055.1988A5 at localhost>,
Wolfgang Schechinger <wolfsc at ibms.sinica.edu.tw> wrote:
>
>In contrary,  I would like to be sure that only one copy enters a cell (I 
>testing a library on eucaryotic cells). What methods for transfection would 
>you recommend? 

Electroporation *tends* to put fewer plasmids per cell, but I htink there 
are still many.

There's a protocol in which you fuse the membranes of your 
plasmid-containing bacteria with your cells for transfection, and then 
dump on a ton of antibiotic.  That's supposed to put a single plasmid per 
cell.  (Actually, of course, many plasmids, but all identical clones.)  I 
believe it's in the Red Book of molecular biology protocols.  I've never 
tried it myself.

>What about multiple stable transfection of individual cells using retrovirus (I 
>currently do that and assume ~one infection per cell)

How many selection markers do you have?  

If you're testing a library, are you planning on re-cloning your library 
into the retrovirus?

Ian 
-- 
    Ian York   (iayork at panix.com)  <http://www.panix.com/~iayork/>
    "-but as he was a York, I am rather inclined to suppose him a
     very respectable Man." -Jane Austen, The History of England




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