Product nameHydroxymethylated DNA Immunoprecipitation (hMeDIP) Kit
See all hydroxymethylated DNA kits
Assay time3h 00m
Species reactivityReacts with: Mouse, Human
DNA methylation is a covalent modification of the cytosine ring at the 5' position, resulting in 5-methylcytosine (5-mC). In somatic cells, 5-mC is found almost exclusively in the context of paired symmetrical methylation of the dinucleotide CpG. The biological important of 5-mC as a major epigenetic modification in phenotype and gene expression has been recognized widely. Quite recently, a novel modified nucleotide, 5-hydroxymethylcytosine (5-hmC), has been detected to be abundant in mouse brain and embryonic stem cells. It is a hydroxylated and methylated form of cytosine. Although it is still uncertain its specific role in epigenetics, it is believed it plays a role in DNA demethylation, chromatin remodeling and gene expression regulation.
Hydroxymethylated DNA Immunoprecipitation (hMeDIP) Kit (ab117134) enables the user to enrich hydroxymethated DNA using an antibody specific to hydroxymethylcytosine (5-hmC) to immunoprecipitate hydroxymethylated genomic DNA hMeDIP. The enriched hydroxymethylated fractions can be used in various downstream applications including PCR (hMeDIP-PCR) and microarrays (hMeDIP-chip) for investigating gene-specific hydroxymethylation in cells or tissues.
Storage instructionsPlease refer to protocols.
Components 24 tests 48 tests 96 tests 10X Wash Buffer 1 x 5ml 1 x 10ml 1 x 20ml 5-hmC Antibody, 0.6 mg/mL 1 x 25µl 1 x 50µl 1 x 100µl 8-Well Assay Strips (with Frame) 3 units 6 units 12 units Adhesive 8-Well Strip Film 3 units 6 units 12 units Antibody Buffer 1 x 4ml 1 x 8ml 1 x 16ml Control DNA, 500 ng/mL 1 x 5µl 1 x 10µl 1 x 20µl Control Primer-Forward, 20 µM 1 x 5µl 1 x 10µl 1 x 20µl Control Primer-Reverse, 20 µM 1 x 5µl 1 x 10µl 1 x 20µl DNA Release Buffer 1 x 7ml 1 x 14ml 1 x 28ml hMeDIP Solution 1 x 3ml 1 x 6ml 1 x 12ml Non-Immune IgG ,0.6 mg/mL 1 x 10µl 1 x 20µl 1 x 40µl Proteinase K, 10 mg/mL 1 x 28µl 1 x 56µl 1 x 112µl
Sensitive detection of gene-specific hydroxymethylation by hMeDIP-QPCR.
Human brain DNA (500 ng) was fragmented to 200-600 bps with a sonicator. The fragmented DNA was used for hydroxymethylated DNA enrichment with ab117134. Eluted DNA was analyzed by real time PCR with primers specifically for OCT4 or GAPDH sequences in the promoter regions. Results show that the promoter region is hydroxymethylated in OCT4 but not in GAPDH.
Selective enrichment of hydroxymethyated DNA with ab117134.
50 pg of unmethylated, methylated, and hydroxymethylated DNA control were spiked into fragmented human genomic DNA (500 ng). hMeDIP was processed using 5-hmC antibody and non-immune IgG included in the kit. Eluted DNA was analyzed by real time PCR using control primers included in the kit to detect the presence of spiked control DNA.
This product has been referenced in:
- Yilmaz-Ozcan S et al. Epigenetic mechanisms underlying the dynamic expression of cancer-testis genes, PAGE2, -2B and SPANX-B, during mesenchymal-to-epithelial transition. PLoS One 9:e107905 (2014). Functional Studies ; Human . Read more (PubMed: 25229454) »