Data

Preparing DNA Samples for Next-Gen Sequencing following MethylCollector Ultra Enrichment

Enriched methylated DNA obtained using MethylCollector Ultra can be used directly in Next-Gen Sequencing sample processing protocols.

If you intend to perform Next-Gen sequencing analysis following MethylCollector Ultra enrichment of methylated DNA, please refer to the MethylCollector Ultra manual and the Illumina ChIP-Seq DNA Sample Prep Kit for instructions on how to prepare your DNA samples for Next-Gen sequencing.

Click on image to enlarge size.

Figure 1:  Next-Gen sequencing data generated using MethylCollector Ultra enriched DNA correlates well with CpG density.

DNA was enriched from 1 µg of human PBMC DNA using Active Motif’s MethylCollector Ultra Kit. Next-Gen sequencing was performed using the Illumina platform to generate 28 million sequence tags. Tags were mapped to generate a whole-genome DNA methylation profile. The image above shows that the enriched regions (purple peaks) correlate well with CpG density except at CpG islands, which should be largely unmethylated.


Click on image to enlarge size.

Figure 2:  Next-Gen sequencing data generated using MethylCollector Ultra enriched DNA is more sensitive and robust than data from other MBD enrichment kits.

DNA was enriched from 1 µg of human PBMC DNA using Active Motif’s MethylCollector Ultra Kit and a competitor’s MBD-based enrichment kit. Next-Gen sequencing was performed using the Illumina platform to generate 28 million sequence tags. Tags were mapped to generate whole-genome DNA methylation profiles. The top purple panel is data produced from MethylCollector Ultra and the orange panel is data from the Competitor M kit. MethylCollector Ultra identified more methylated regions and the peaks had greater intensity than peaks identified from the competitor kit.


Click on image to enlarge size.

Figure 3:  Next-Gen sequencing data generated using MethylCollector Ultra detects methylation at CpG shores.

DNA was enriched from 1 µg of human PBMC DNA using Active Motif’s MethylCollector Ultra Kit. Next-Gen sequencing was performed using the Illumina platform to generate 28 million sequence tags. Tags were mapped to generate a whole-genome DNA methylation profile. The image above shows one of many examples of DNA methylation being detected at CpG shores rather than in the CpG island itself. This data agrees with recent findings showing that a large portion of methylated sites occur in these CpG shore regions that are adjacent to CpG islands.