Methylation of Sfmbt2 CpG Island. A. Five amplicons representing 140 CpG sites across the transcriptional start sites for both the sense and antisense transcripts at the Sfmbt2 locus were queried for methylation in sperm and oocyte genomic DNA by bisulfite mutagenesis. Two amplicons, at the 5' and 3' extremes of the CpG island, Me3 and CG3, were also examined in C57BL6 X Castaneus F1 placenta genomic DNA to determine parental allele methylation. Black circles represent CpGs that survived bisulfite conversion; open circles represent CpGs that were converted to TG; red circles represent asymmetric cytosines in oocyte genomic DNA that survived conversion in most clones; grey circles represent cytosines in a CpG context that were methylated on only the bottom strand in oocyte genomic DNA. Each line represents a unique clone; duplicate clones were not scored. Bisulfite conversion was approximately 95%. The apparent two CpG sites in CG1 are only separated by a single base pair, and are therefore assumed to be a single site. B. Relative location of bisulfite amplicons within CpG island. Hatched boxes represent amplicons for which only one strand was queried; solid box (CG3) was analysed on both the top and bottom strands. Hash marks indicate the relative location of the methylated cytosines in oocyte genomic DNA; black = CpG, red = asymmetric. Of 25 methylated cytosines, 18 were confirmed to be asymmetric, including the four CpG sites in CG3 illustrated in panel A.