Need help with non-rad Southerns

Lauryl Maiya J Nutter lmjnutte at acs5.acs.ucalgary.ca
Mon Feb 5 19:40:54 EST 1996


Robert Pierce (rpierce at u.washington.edu) wrote:
: I have gotten my hands on some dig-dUTP and would like to 
: give a non-radioactive Southern a try, but am somewhat baffled by the 
: question as to the best membrane and detection system to employ.  Any 
: helpful hints would be greatly appreciated.  Thanks in advance.

: Rob

The membrane you choose and the detection system you use depends
on the sensitivity that you need for your Southern.  For a single
copy gene from Drosophila DNA I have been using either
Photogene membrane from GIBCO/BRL or Hybond N from Amersham along
with the B.M. CSPD DIG chemiluminescent detection protocol.  For
more abundant DNAs (eg. PCR reactions, colony lifts, plasmid
screens) I have used the same membranes as well as Hybond N+ from
Amersham (the folks upstairs routinely use Zetaprobe membranes
with bacterial genomic DNA) and NBT/X-phos colorimetric
detection.

Hope this helps.

Good luck!

Lauryl Nutter
University of Calgary
lmjnutte at acs.ucalgary.ca



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