A tag-based approach for high-throughput analysis of CCWGG methylation.
Non-CpG methylation occurring in the context of CNG sequences is found in plants at a large number of genomic loci. However, there is still little information available about non-CpG methylation in mammals. Efficient methods that would allow detection of scarcely localized methylated sites in small quantities of DNA are required to elucidate the biological role of non-CpG methylation in both plants and animals. In this study, we tested a new whole genome approach to identify sites of CCWGG methylation (W is A or T), a particular case of CNG methylation, in genomic DNA. This technique is based on digestion of DNAs with methylation-sensitive restriction endonucleases EcoRII-C and AjnI. Short DNAs flanking methylated CCWGG sites (tags) are selectively purified and assembled in tandem arrays of up to nine tags. This allows high-throughput sequencing of tags, identification of flanking regions, and their exact positions in the genome. In this study, we tested specificity and efficiency of the approach.
Denisova
Oksana V
OV
Institute of Protein Research, Russian Academy of Sciences, 142290 Pushchino, Russian Federation.
Chernov
Andrei V
AV
Koledachkina
Tatyana Y
TY
Matvienko
Nicholas I
NI
D000818:Animals; D001483:Base Sequence; D019175:DNA Methylation; D017931:DNA Primers / Q000235:genetics*; D015257:DNA-Cytosine Methylases / Q000235:genetics* / Q000378:metabolism; D015252:Deoxyribonucleases, Type II Site-Specific / Q000235:genetics* / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D023281:Genomics / Q000379:methods*; D006801:Humans; D015203:Reproducibility of Results
D017931:DNA Primers; C029528:DNA modification methylase EcoRII; D015257:DNA-Cytosine Methylases; C046545:CCWGG-specific type II deoxyribonucleases; D015252:Deoxyribonucleases, Type II Site-Specific