hplc of plasmids/small DNA fragments?

Brad Thomas bthomas at gmm.gen.emory.edu
Tue Oct 31 14:20:29 EST 1995


Waters Corp. has a fast anion exchanger that is useful for analysis and 
purification of DNA:  its called Gen-Pak FAX.  Other companies have 
these as well, example Perkin Elmer.  In general they use 2 to 3 micron 
nonporous particles.  These give much better resolution than the various 
porous ion exchangers that are marketed for DNA analysis and 
purification.  The Gen-Pak has a higher capacity than most and is 
probably better suited for purification than its competitors.  
Methodology has been developed for separation of DNA on nonporous 
divinylbenzene reversed phase columns.  Resolution looks very good but I 
haven't tried it myself.  The advantage here seems to be that the DNA 
comes out in much lower salt than you have to deal with when using anion 
exchangers. 
Using anion exchange (or reversed phase), you can count on separations 
taking 10-30 minutes with resolution equal to agarose gels for DNA from 
<20 to around 10 kb.  Usable separations can be gotten from 10-30 kb.  
You can also get good separations between double-strand and single 
strand, supercoiled and nicked circles etc. 





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