About Quest 5-hmC™ DNA Enrichment Kit
While the importance of DNA methylation in epigenetic regulation is well established, the biological role of hydroxymethylation remains elusive. The “sixth base”, 5-hydroxymethylcytosine (5-hmC), has been detected in the DNA of embryonic stem cells and other cell types. Brain tissue DNA contains the highest levels of 5-hmC. Recent work suggests that 5-hmC may function in gene regulation and may be involved as an intermediate in active demethylation of 5-methylcytosine (5-mC).
The Quest 5-hmC™ DNA Enrichment Kit features J-base binding protein (JBP) for the specific enrichment of 5-hmC containing DNA. The consolidated workflow makes the procedure reliable for robust analysis of multiple samples. Simply glucosylate the input DNA, add JBP Capture MagBeads, and then wash and elute the enriched 5-hmC DNA.
Specifications
| Equipment | Microcentrifuge, heat block, incubator, and a rotator/rocker are required. A magnetic stand is recommended, but this kit includes magnetic rods if such a stand is unavailable. A thermal cycler will be necessary to perform PCR. |
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| Sample Sources | This protocol is developed for processing fragmented genomic DNA as the input material. DNA should be free of enzymatic inhibitors and can be in water, TE, or low salt buffer. |
| Input | 5-4,000 ng DNA |
| Purity | Enriched 5-hmC DNA is of suitable purity for a wide variety of downstream applications including PCR, qPCR, Next-Gen sequencing, whole genome amplification, arrays, and more. |
| 5-hmC Glucosyltransferase | One unit (1 U) of recombinant 5-hmC Glucosyltransferase (5-hmC GT) is sufficient to glycosylate 1 μg of 5-hmC containing DNA in 1 hour at 37 °C. |
