Before cloning, a DNA fragment needs double digestion to create two cohesive ends, thus two different enzymes are required. If the best performance of these two enzymes varies in different buffer system, what is the best solution to circumvent this problem?
You need to check the possibility of using a common buffer if the enzymes are 50-100% compatible (active). Otherwise sequential digestion need to be done.
https://www.neb.com/tools-and-resources/usage-guidelines/double-digests
nowadays, many RE cut in cut smart buffer from NEB..;these are highly concentrated enzyme . they usualy cut nicely.
cuts at the end of PCR product is always of concern, be carefull to add extra nt , i usually add 6 ..and if I experience trouble I cut OVN in a culture incubator or pcr machine
Thank you very much your kind suggestions are very helpful.
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