65 and 64 separation

Hiranya Roychowdhury hroychow at NMSU.EDU
Mon Aug 8 12:09:50 EST 1994


On 5 Aug 1994, jinsong sheng wrote:

> 
> Dear Netters:
> 
> Any suggestions to separate a 65 and 64 kd proteins on SDS -PAGE?  Like 
> percentage of gels, gradient or buffer systems?  
> 
> Many thanks in advance.
> 
> Jinsong Sheng.
> 
> 
The % of the gel can be varied and the separation tried out. I would try
three different gradients: 10-15%, 10-12% and 8-12%. However, since the
separation is often dependent on the type of protein (state of conjugation
etc.), one can also obtain reasonable separation by just an isocratic gel.
Again, the % T has to be chosen by trial-and-error.

If this simple step does not help then one can manipulate the %C (ie.
instead of a regular 4 or 4.5%C).

Tricine buffer system (instead of glycine), which also employs a higher
%C, has been known to separate smaller polypeptides (below 12 kDa) rather
efficiently. I am not sure if it will work for moderately higher mol. wt.
polypeptides. Worth a try.

Good Luck.

Hiranya . 







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