BclI methylation sensitivity - yes or no or what?
tk at mit.edu
Tue Mar 14 15:48:51 EST 2006
"Wolfgang Schechinger" <novalidaddress at nurfuerspam.de> writes:
> BclI (SpeI) is reported to be dam methylation sensitive (see eg NEB and
> Fermentas catalogues)
> just was cutting pLNCX into 2 fragments with BclI (Bcl one). As the
> plasmid was accidentally prepared in DH5alpha, actually BclI shouldn't
> have cut at all. On the gel, the digest seemed to be complete. The
> reaction was set up as usual (5 Units enzyme, 1hrs 50degC, Rxn vol 50
> What went right? Your comments are appreciated!
If you look here: http://rebase.neb.com/cgi-bin/msget?BclI
you'll see that BclI will cut hemimethylated DNA pretty effectively.
If your DNA was harvested from a log phase culture, much of it
probably was hemimethylated rather than fully methylated.
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