EpiJET 5-hmC Enrichment Kit

The Thermo Scientific EpiJET 5-hmC Enrichment Kit enables the enrichment of 5-hmC containing DNA over unmodified DNA and 5-mC DNA. The kit takes advantage of the 5-hmC Modifying Enzyme, which is formulated for highly specific and efficient modification of 5-hmC present in CpG dinucleotides of DNA and is deficient for any activity over unmodified or methylated cytosines. The enzyme adds a short linker molecule to 5-hmC, to which biotin is conjugated in the following chemical modification step. The resulting modified DNA is separated from unmodified DNA using streptavidin-coated magnetic beads.

More than 100-fold enrichment of the 5-hmC modified DNA over unmodified and 5-mC containing DNA and at least 20% recovery of the input 5-hmC DNA can be achieved. For performance evaluation of the kit, specific DNA controls (unmodified and 5-hmC DNA) are included.

Highlights

•     Specific – no activity over unmodified or methylated cytosines
•     Versatile – efficient enrichment of single and multiple 5-hmC targets
•     Robust – consistent yield and enrichment
•     Fast – procedure completed in 3 hours
•     Pure – enriched DNA suitable for downstream applications

Applications

•     PCR
•     qPCR
•     Sequencing
•     Microarrays
•     NGS

500 ng of ultrasound fragmented human brain (high levels of 5-hmC) or blood DNA (low levels of 5-hmC) was enriched using the EpiJET 5-hmC Enrichment Kit. Resulting yields of different loci were estimated using qPCR as a fraction of the input DNA. Higher enrichment levels of brain DNA loci in comparison to blood DNA demonstrate the specificity of the kit for 5-hmC.

Includes

•     Box 1/2

        5-hmC Modifying Enzyme
        4X Enzyme Reaction Buffer
        10X Biotin Conjugation Buffer
        Biotin Reagent
        Primer Mix for Control DNA (unmodified), 5 µM
        Primer Mix for 5-hmC Control DNA, 5 µM
        5-hmC Control DNA, 10 ng/µL
        Control DNA (unmodified), 0.5 µg/µL

•     Box 2/2

        Streptavidin Magnetic Beads
        Streptavidin Binding Buffer
        Streptavidin Wash Buffer 1
        Streptavidin Wash Buffer 2
        Water, nuclease-free
        Column Binding Buffer
        Column Wash Buffer (concentrated)
        Elution Buffer
        DNA Purification Micro Columns and Collection Tubes