acid phenol extractions

brett brett at BORCIM.WUSTL.EDU
Tue Feb 18 11:03:28 EST 1997


>Hi all,
>
>I just tried an acid phenol extraction of a plasmid prep in order to get
>rid of nicked plasmid. I'm later doing exo III deletions. I equilibrated
>the phenol to pH 4.0 (or perhaps even a little bit lower) and did the
>extractions on about 10 ug of plasmid DNA. Even though the manual said you
>should expect to see a white band at the interphase, I didn't really see
>one. After three extractions, and a final CHCl3:IAA extraction, I tried to
>precipitate the DNA. The yield is abysmal. I pretty much lost all my DNA.
>I had run some of the plasmid before hand, so I knew there was some nicked
>DNA for sure, but there also was plenty of ccc plasmid. Has anyone
>done this protocol before ? any pointers you can give me ?
>
>Ed

The only advise I can give is to cesium band your DNA, or try and work with
the nicked stuff.


Brett Lindenbach
    
Program in Immunology                              
Washington University - St Louis                  
brett at borcim.wustl.edu                             




More information about the Methods mailing list