Methylated DNA Interfering with Restriction Digestion - Restriction digestion problems (Oct/16/2006 )

Hi All,

I'm wondering what to do to solve the problem of getting my plasmid to digest with ApaI when the target DNA is methylated. Since methylation is cell dependent, can I just re-transform DNA into a methylation negative strain and do preps to obtain unmethylated DNA? Any help would be great. Thanks in advance.

Yeah, I am pretty sure that a strain like DH5alpha or XL-1 blue will be fine. Just purify your DNA from a strain that doesn't methylate and you should be able to digest your DNA no problem.

Both DH5a and XL-1 Blue are dam+ and dcm+ and will methylate the DNA. Look for the NEB strains, ER2925, for example. See
http://www.neb.com/nebecomm/tech_reference...hylated_dna.asp

Thank you both for your input. I was able to use BL21 strain (which is dcm-) to solve the problem. It's amazing how you learn something new every day in working in science.