PFGE with LMP?

[Andrei Popov]

STACY A CIUFO wrote:
> 
> Hello everyone,
> 
> Has anyone ever used low melting point agarose for
> pulsed field gel electrophoresis?
> 
> Regular agarose works well, but LMP gives terrible
> results.  If any of you have experience in resolving
> and isolating large DNA fragments from each other,
> please let me know how.
> 

hello Stacy, 

the majority of the YAC-libraries were constructed from 
DNA-fragments separated on LMA gels. Yes, the resolution on 
LMA gels is a bit worse, but it sould not be "terrible".
Try to change the brand of the LMA; increase the percentage from (I 
suppose) 1% to 1.5%; decrease the voltage/increase the time of the run; 
and the last, but probably most important, set the temp of the cooling 
bath to 3.5C. There are other tricks, for example I found that 
increasing the thickness of the gel (whether LMA or not) makes the DNA
bands to run under a funny angle.


Best wishes

Andrei