Differential Display

Daniel Kim dkim at nmsu.edu
Sat Apr 22 10:12:39 EST 1995


Re:  Not getting bands in differential display.

I have used the differential display technique in the past, and found 
that I got better results if I used a lot of enzyme.  I had as much as 2 
units of Taq polymerase per reaction (50 microliter).  You can get away 
with 1 unit, but 2 is more reliable.

I don't know why this was the case, but it made the reactions more 
robust.  It might be that there are more active sites of polymerization 
in a differential display reaction, and so more enzyme is needed to 
support it.  Remember that the standard single-product PCR reaction 
starts off making very little DNA in the initial reaction cycles and 
works its way up, making one band.  The differential display (and other 
random-primed methods) will start off making many different DNA 
molecules, and amplify each of them.  You should need a much larger 
amount of enzyme to support it.

If there are "responsible opposing points of view", I'd like to hear 
them. I don't really know enough to make a solid case for my opinion here.

Daniel Kim



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