DNA methylation sites

[Andrew Hobbs]

Brian H. Taylor (BRIAN at BIO.TAMU.EDU) wrote:
: 
:     I'm wondering if anyone has ever ruled out cytosine methylation at
: GC dinucleotides (as opposed to CG and CXG) or, for that matter,
: CXXG. I'm familiar with the Gruenbaum et al. Nature paper from 1981
: (Vol. 292, 860-862), where nearest neighbor analysis was used to show
: a high level of methylation at CG and CXG.  One nonsymmetrical
: arrangement, CAT, was tested and was found to be <4% methylated, but
: I don't know if that means it was below their limits of detection or
: they could see almost 4% methylation at those sites.  I don't think
: their approach would have tested for GC symmetry.  The reason I am
: asking is that I occasionally encounter cases of possible incomplete
: digestion of restriction sites in genomic plant DNA with enzymes
: like HindIII (recognition sequence AAGCTT).  This should not be
: methylated by the usual rules, but if either GC or CXXG methylation
: occurs it would block digestion.  Appropriate controls indicate that
: I don't have a problem with dirty DNA or bad enzyme or any of the
: usual trivial explanations.  Any additional information on
: alternative methlation sites will be appreciated.
: 
: 
: 
: 
: 
: Brian H. Taylor                         *   Ad hoc loc and
: Dept. of Biology, Texas A&M University  *   quid pro quo,
: College Station, TX  77843-3258         *   so little time,
:                                         *   so much to know!
: Phone:  (409) 845-7754                  *
: FAX:    (409) 845-2891                  *   Jeremy Hilary Boob
:                                         *   (The Nowhere Man)

What you are finding is typical for plant DNA.  Methylation of Cytosines
can reach close to 100% in plant genomic DNA.  (Not just in CG
dinucleotides).  Hence when using restriction enzymes to digest plant
DNA it is best to avoid enzymes which are inhibited by methyl-C at any
position in their recognition site.

Andrew Hobbs
Dept of Biochemistry
University of Western Australia

andrewh at uniwa.uwa.edu.au