Lambda DNA resists digestion

[Thomas Newman]

In article <396aas$ki1 at news.service.uci.edu>, rmauk at crick.bio.uci.edu (Rob
Mauk) wrote:

> 
> I have isolated lambda gt10 DNA by PEG precipitation followed by organic 
> extraction and ethanol precipitation, and find that it is almost 
> completely resistant to digestion by EcoRI and all other enzymes that I 
> have tested.  Proteinase K treatment did not help, nor did additional 
> extractions and precipitations.  Plasmid DNA digests normally with these 
> enzymes, but if the plasmid is mixed with the lambda DNA, it is rendered 
> undigestable.  Any comments or suggestions regarding the nature of this 
> contaminant, or solutions to the problem would be greatly appreciated.  
> Thank you.

I have found in many cases that the addition of spermidine to a final
concentration of 4 mM in the restriction buffer will allow the enzymes to
work.  It is an easy test!!

Tom Newman
Assistant Prof.
MSU-DOE Plant Research Laboratory