Identification of 5-methylcytosine in complex genomes.
Cytosine methylation is attracting new attention for regulatory roles in gene expression and there is an increasing interest in detecting, at a single-base resolution, any 5-methylcytosine in genes from complex genomes. Differential base modification by chemicals followed by PCR-based genomic sequencing procedures can provide the resolution, sensitivity, and specificity required for such a goal. The various methods available are not devoid of artifacts but if used carefully and in combination, very reliable information can be obtained. We compare the methods using bisulfite and conventional PCR with those using either hydrazine or potassium permanganate and ligation-mediated PCR and provide a step-by-step description of the corresponding procedures.
Thomassin
H
H
Institut Jacques Monod du CNRS, Université Paris 7, Tour 43, 2 Place Jussieu, Paris Cedex 05, 75251, France.
Oakeley
E J
EJ
Grange
T
T
D044503:5-Methylcytosine; D002467:Cell Nucleus / Q000737:chemistry* / Q000235:genetics; D003596:Cytosine / Q000031:analogs & derivatives* / Q000032:analysis; D019175:DNA Methylation; D016678:Genome; D008967:Molecular Biology / Q000379:methods*
D044503:5-Methylcytosine; D003596:Cytosine