I am in need of doing partial digestion for some of my cloning; however, I have fastdigist enzymes aviable and I can not find a suitable protocol for achieving partial digests with these enzymes. Has anyone done partial digests with FastDigest Enzymes before? If so what protocol was used?
You can do this by incubating them 1/10 of the time than the protocol says. If I were you I would fix the protein concentration and just play with the DNA concentration to get a good chance to get the result you want quick.
Use reaction temperature 20'C and incubation time of max. 15 min . We have used this protocol with FastDigest enzymes.
Assuming you have plenty of DNA I would just set up a dilution series with constant DNA and 2X dilution of enzyme for each tube. Incubate all for a standard time / temperature and analyze by gel to identify appropriate conditions.
Thanks for all the suggestions. I found a two-fold serial dilution of the enzyme, with 1-2min incubation time works well with FastDigest. They are very efficient so you have to modify both enzyme and incubation parameters.
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