Hello!
Has anyone ever performed partial digestion of DNA, I try to cut DNA this way, but it is hard to match the conditions. Maybe there is some clever method to make it efficiently.
Regards
Tomek
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tfitzwater
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Posted 15 December 2004 - 04:41 PM
There are some old tricks.
If you add ethidium bromide to the restriction digest, the enzyme will make a single cut and stop. I don't remember the exact concentration of EtBr required.
Another option dates back to the old days of cesium gradient purification of plasmid DNA. The preferential initial cut by a restriction enzyme can vary depending upon whether or not the DNA has been run over a cesium gradient and then phenol:chloroform extracted. The same might be true of DNA that has been prepared by alkaline lysis and then plus or minus purification by one of the numerous commercial kits.
If you add ethidium bromide to the restriction digest, the enzyme will make a single cut and stop. I don't remember the exact concentration of EtBr required.
Another option dates back to the old days of cesium gradient purification of plasmid DNA. The preferential initial cut by a restriction enzyme can vary depending upon whether or not the DNA has been run over a cesium gradient and then phenol:chloroform extracted. The same might be true of DNA that has been prepared by alkaline lysis and then plus or minus purification by one of the numerous commercial kits.
