Combining gel filtration and electrophoresis

Austin P. So (Hae-Jin) nobody at nowhere.com
Wed Feb 1 18:11:59 EST 2006


Daniel Käsmayr wrote:

> What I would like to have is a system that uses electric charge to  
> drive particles, but also a matrix that retards smaller particles  much 
> more than larger ones; resulting in something like an inverse- PAGE, 
> where larger particles migrate faster than smaller ones.
> 
> The substances that need to be characterized are star polymers /  brush 
> polymers that have multiple functional groups (amine or  carboxylic 
> acids) which are used to create peptide bonds to make  conjugates with 
> DNA and/or Proteins.
> 
> What would also help is some sort of modification I could do to PAGE  
> that would help me get my star-shaped polymers (and their conjugates  
> with proteins or DNA) into the gel. Right now it seems like I either  do 
> not get them to penetrate the gel matrix (I see only some residues  on 
> the gel pocket's outside and just a few mm inside the gel - these  are 
> 3% gels already and ver un-handlable) or they are not stainable  with a 
> silver stain…
>  From my syntheses I know the polymers can be analyzed by GPC/SEC -  but 
> I don't have enough material for this anymore - DNA is very  expensive 
> if you are talking about mg amounts.

Hmmmm...I don't get it since there is no apparent reason (at least to 
me), why you need to fractionate things in the reverse manner...do you 
need to retain the larger products? Is there a reason why 
membrane-fitration at a specific MWCO wouldn't work?

BTW, DNA is probably the cheapest thing to obtain in mg quantities.

Nonetheless, one way that I can think of is a method that was developed 
by a professor out here at UBC:

http://www3.interscience.wiley.com/cgi-bin/abstract/109861279/ABSTRACT?CRETRY=1&SRETRY=0

Basically, they exploit the fact that there is a non-linear relationship 
between size and electrophoretic velocity, so that basically you can 
encourage the migration of large fragments of DNA.

Austin


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