EpiXplore Meth-Seq DNA Enrichment Kit

The EpiXplore Meth-Seq DNA Enrichment Kit is a methylated DNA enrichment and library prep kit. This is a product for separating the methylated and unmethylated fractions of DNA, and generating sequencing-ready Illumina® libraries from sheared genomic DNA. This kit utilizes his-tagged MBD2 protein and Capturem^TM Nanoprep columns to separate methylated DNA from unmethylated DNA in ~2 hours, and employs our DNA SMART^TM technology to generate indexed meth-seq libraries suitable for NGS on Illumina sequencing platforms. Sequencing libraries are generated without the need for enzymatic cleanup or additional adapter ligations. All necessary reagents for producing meth-seq libraries are provided in this kit, including primer sets that contain Illumina index sequences. For more information on the indexes supplied in each kit, please refer to the relevant Certificates of Analysis.

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Flowchart for separating methylated and unmethylated fractions of DNA

Flowchart for separating methylated and unmethylated fractions of DNA. In the enrichment portion of the protocol, sheared DNA is incubated with his-tagged MBD2 protein for 1 hr, which allows the methylated DNA fraction to bind to the protein. The solution is then added to the Capturem nanoprep column, which binds the his-tagged protein and thus the methylated DNA. Subsequent washes with the Unmethylated Elution Buffer remove the unmethylated DNA. The methylated DNA is then eluted in a high-salt buffer. After cleanup, this enriched DNA can be used as the input for library preparation.

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Flowchart for DNA-SMART ChIP-Seq technology

Flowchart for DNA-SMART ChIP-Seq technology. This single-tube workflow allows users to generate Illumina-compatible libraries for meth-seq experiments. After library size selection and purification, the total time from input DNA to meth-seq library is approximately four hours. DNA SMART technology eliminates the need for an adapter ligation step and associated cleanup, reducing the loss of limited input DNA.

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Distinct methylation patterns are observed in aligned NGS data of libraries generated from methylated and unmethylated DNA fractions

Distinct methylation patterns are observed in aligned NGS data of libraries generated from methylated and unmethylated DNA fractions. A representative region of Arabidopsis chromosome 3 (outlined in the green box) shows the peaks identified in the DNA enriched for methylated and unmethylated fractions. The methylated regions are outlined in red boxes and the unmethylated regions are outlined in blue boxes. The amount of initial input DNA is shown along the left-hand side of the alignments. The data show good concordance with the publicly available bisulfite sequencing data. Data were visualized using the Integrative Genomics Viewer (Broad Institute). Public data was obtained from GEO accession GSM980986; Stroud et al., 2013.