Help! I made a mistake

David F. Spencer dspencer at is.dal.ca
Fri Nov 7 11:53:50 EST 1997


In article <peng.36.190.34633C9E at postbox.acs.ohio-state.edu>,
peng.36 at postbox.acs.ohio-state.edu (p.J) wrote:

> Hi, neters:
>        I made a wrong SSC solution.  It sould be sodium chloride and
Trisodium 
> citrate.  But I made it with sodium chloride and Tris base.  The worst thing 
> is that I used this SSC as a transfer buffer to transfer the DNA from 1% 
> agrose gel to membrane when I did Southern.  Now, I do not know what I should 
> do, trash the membrane or contine.  Any body have any suggestions?  Thanks a 
> lot.P. J

I wouldn't worry. I'm assuming you're using nylon rather than
nitrocellulose. What you've done is a quasi-alkaline transfer. I use the
standard alkaline transfer method which is with 0.4M NaOH alone, whereas
straight Tris (i.e. unneutralized) can't have a pH higher than about 11 (1M
Tris is about 11.2).  You may actually find that your transfer worked
better than usual!

Dave

--------------------------------------------
David F. Spencer, PhD
Dept. Of Biochemistry
Dalhousie University
Halifax, Nova Scotia
Canada

dspencer at is.dal.ca
dspencer at rsu.biochem.dal.ca



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