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MethylCap kit

Catalog Number
48 rxns

The MethylCap assay is based on the affinity purification of methylated DNA using a MBD (Methylbinding domain) protein . This procedure has first been optimized on the SX-8G IP-Star Automated System ([page product SX-8G IP-Star]. Afterwards, this method has been adapted to a manual protocol, which has been inspired by the same reproducibility standards provided by the automated MethylCap assay.

  • Characteristics
    • Allows for fractionation of methylated DNA by CpG density
    • Compatible with next generation sequencing
    • Magnetic beads allow for fast & sensitive capture of methylated DNA
    • Includes control primer pairs for the assessment of the capture efficiency

    Methylation of CpG dinucleotides is generally associated with epigenetic silencing of transcription and is maintained through cellular division. Multiple CpG sequences are rare in mammalian genomes, but frequently occur at the transcriptional start site of active genes, with most clusters of promoter CpGs being hypomethylated (Kangaspeska, 2008).

    The Diagenode H6-GST-MBD fusion protein of the MethylCap kit has been extensively validated. It consists of the methyl binding domain (MBD) of human MeCP2, as a C-terminal fusion with Glutathione-S-transferase (GST) containing an N-terminal His6-tag. The H6-GST-MBD fusion protein can be used to specifically isolate DNA containing methylated CpGs. See overview below.

    MBD-sequencing results have been validated by bisulfite sequencing

    Figure 1. Using the MBD approach, two methylated regions were detected in different elution fractions according to their methylated CpG density (A). Low, Medium and High refer to the sequenced DNA from different elution fractions with increasing salt concentration. Methylated patterns of these two different methylated regions were validated by bisulfite conversion assay (B).

  •  Documents
    MethylCap kit MANUAL
    This MethylCap kit includes the methylCap protein (Cat. No. mbd-001-100) which has been extensive...
    Datasheet MethylCap Protein DATASHEET
    This MethylCap protein has been extensively validated for specific isolation of DNA containing me...
    DNA Methylation Solutions BROCHURE
    Complete solution for DNA methylation studies! Whether you are experienced or new to th...
  •  Safety sheets
    MethylCap kit SDS GB en Download
    MethylCap kit SDS US en Download
    MethylCap kit SDS BE nl Download
    MethylCap kit SDS BE fr Download
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    MethylCap kit SDS ES es Download
    MethylCap kit SDS DE de Download
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  •  Publications

    How to properly cite this product in your work

    Diagenode strongly recommends using this: MethylCap kit (Diagenode Cat# C02020010). Click here to copy to clipboard.

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    SMaSH: Sample matching using SNPs in humans.
    Westphal M, Frankhouser D, Sonzone C, Shields PG, Yan P, Bundschuh R
    BACKGROUND: Inadvertent sample swaps are a real threat to data quality in any medium to large scale omics studies. While matches between samples from the same individual can in principle be identified from a few well characterized single nucleotide polymorphisms (SNPs), omics data types often only provide low to mod...

    Increased presence and differential molecular imprinting of transit amplifying cells in psoriasis.
    Witte K, Jürchott K, Christou D, Hecht J, Salinas G, Krüger U, Klein O, Kokolakis G, Witte-Händel E, Mössner R, Volk HD, Wolk K, Sabat R
    Psoriasis is a very common chronic inflammatory skin disease characterized by epidermal thickening and scaling resulting from keratinocyte hyperproliferation and impaired differentiation. Pathomechanistic studies in psoriasis are often limited by using whole skin tissue biopsies, neglecting their stratification and ...

    Plasticity of histone modifications around Cidea and Cidec genes with secondary bile in the amelioration of developmentally-programmed hepatic steatosis.
    Urmi JF, Itoh H, Muramatsu-Kato K, Kohmura-Kobayashi Y, Hariya N, Jain D, Tamura N, Uchida T, Suzuki K, Ogawa Y, Shiraki N, Mochizuki K, Kubota T, Kanayama N
    We recently reported that a treatment with tauroursodeoxycholic acid (TUDCA), a secondary bile acid, improved developmentally-deteriorated hepatic steatosis in an undernourishment (UN, 40% caloric restriction) in utero mouse model after a postnatal high-fat diet (HFD). We performed a microarray analysis and focused ...

    DNA Methylation and Regulatory Elements during Chicken Germline Stem Cell Differentiation.
    He Y, Zuo Q, Edwards J, Zhao K, Lei J, Cai W, Nie Q, Li B, Song J
    The production of germ cells in vitro would open important new avenues for stem biology and human medicine, but the mechanisms of germ cell differentiation are not well understood. The chicken, as a great model for embryology and development, was used in this study to help us explore its regulatory mechanisms. ...

    Antioxydation And Cell Migration Genes Are Identified as Potential Therapeutic Targets in Basal-Like and BRCA1 Mutated Breast Cancer Cell Lines
    Privat M. et al.
    Basal-like breast cancers are among the most aggressive cancers and effective targeted therapies are still missing. In order to identify new therapeutic targets, we performed Methyl-Seq and RNA-Seq of 10 breast cancer cell lines with different phenotypes. We confirmed that breast cancer subtypes cluster the RNA-Seq ...

    Gene body methylation is involved in coarse genome-wide adjustment of gene expression during acclimatization
    Groves Dixon1, Yi Liao1, Line K. Bay2 and Mikhail V. Matz
    Gene body methylation (GBM) is a taxonomically widespread epigenetic modification of the DNA the function of which remains unclear 1,2. GBM is bimodally distributed among genes: it is high in ubiquitously expressed housekeeping genes and low in context-dependent inducible genes 2,3, and it has been hypothesized that...

    Methylome analysis of extreme chemoresponsive patients identifies novel markers of platinum sensitivity in high-grade serous ovarian cancer
    Tomar T. et al.
    BACKGROUND: Despite an early response to platinum-based chemotherapy in advanced stage high-grade serous ovarian cancer (HGSOC), the majority of patients will relapse with drug-resistant disease. Aberrant epigenetic alterations like DNA methylation are common in HGSOC. Differences in DNA methylation are associate...

    ALDH1A3 is epigenetically regulated during melanocyte transformation and is a target for melanoma treatment
    Pérez-Alea M. et al.
    Despite the promising targeted and immune-based interventions in melanoma treatment, long-lasting responses are limited. Melanoma cells present an aberrant redox state that leads to the production of toxic aldehydes that must be converted into less reactive molecules. Targeting the detoxification machinery constitut...

    Male fertility status is associated with DNA methylation signatures in sperm and transcriptomic profiles of bovine preimplantation embryos
    Kropp J. et al.
    BACKGROUND: Infertility in dairy cattle is a concern where reduced fertilization rates and high embryonic loss are contributing factors. Studies of the paternal contribution to reproductive performance are limited. However, recent discoveries have shown that, in addition to DNA, sperm delivers transcription facto...

    Comparative analysis of MBD-seq and MeDIP-seq and estimation of gene expression changes in a rodent model of schizophrenia
    Neary J.L. et al.
    We conducted a comparative study of multiplexed affinity enrichment sequence methodologies (MBD-seq and MeDIP-seq) in a rodent model of schizophrenia, induced by in utero methylazoxymethanol acetate (MAM) exposure. We also examined related gene expression changes using a pooled sample approach. MBD-seq and MeDIP-seq...

    Conserved effect of aging on DNA methylation and association with EZH2 polycomb protein in mice and humans.
    Mozhui K. and Pandey A.K.
    In humans, DNA methylation at specific CpG sites can be used to estimate the 'epigenetic clock', a biomarker of aging and health. The mechanisms that regulate the aging epigenome and level of conservation are not entirely clear. We performed affinity-based enrichment with methyl-CpG binding domain protein followed b...

    Epigenetic sampling effects: nephrectomy modifies the clear cell renal cell cancer methylome
    Van Neste C. et al.
    Purpose Currently, it is unclear to what extent sampling procedures affect the epigenome. Here, this phenomenon was evaluated by studying the impact of artery ligation on DNA methylation in clear cell renal cancer. Methods DNA methylation profiles between vascularised tumour biopsy samples and devascularise...

    Determinants of orofacial clefting II: Effects of 5-Aza-2′-deoxycytidine on gene methylation during development of the first branchial arch
    Seelan R.S. et al.
    Defects in development of the secondary palate, which arise from the embryonic first branchial arch (1-BA), can cause cleft palate (CP). Administration of 5-Aza-2′-deoxycytidine (AzaD), a demethylating agent, to pregnant mice on gestational day 9.5 resulted in complete penetrance of CP in fetuses. Several gene...

    Comparative DNA Methylation and Gene Expression Analysis Identifies Novel Genes for Structural Congenital Heart Diseases
    Grunert M et al.
    Aims For the majority of congenital heart diseases (CHDs), the full complexity of the causative molecular network, which is driven by genetic, epigenetic and environmental factors, is yet to be elucidated. Epigenetic alterations are suggested to play a pivotal role in modulating the phenotypic expression of CHDs a...

    Molecular and epigenetic features of melanomas and tumor immune microenvironment linked to durable remission to ipilimumab - based immunotherapy in metastatic patients
    Seremet T et al.
    Background Ipilimumab (Ipi) improves the survival of advanced melanoma patients with an incremental long-term benefit in 10–15 % of patients. A tumor signature that correlates with this survival benefit could help optimizing individualized treatment strategies. Methods Freshly frozen melanoma met...

    Integrative epigenomic analysis reveals unique epigenetic signatures involved in unipotency of mouse female germline stem cells
    Zhang XL et al.
    Background Germline stem cells play an essential role in establishing the fertility of an organism. Although extensively characterized, the regulatory mechanisms that govern the fundamental properties of mammalian female germline stem cells remain poorly understood. Results We generate genome-wide profiles ...

    RAB25 expression is epigenetically downregulated in oral and oropharyngeal squamous cell carcinoma with lymph node metastasis
    Clausen MJ et al.
    Oral and oropharyngeal squamous cell carcinoma (OOSCC) have a low survival rate, mainly due to metastasis to the regional lymph nodes. For optimal treatment of these metastases, a neck dissection is required; however, inaccurate detection methods results in under- and over-treatment. New DNA prognostic methylation b...

    Genic DNA methylation drives codon bias in stony corals
    Dixon G et al.
    Gene body methylation (gbM) is an ancestral and widespread feature in Eukarya, yet its adaptive value and evolutionary implications remain unresolved. The occurrence of gbM within protein coding sequences is particularly puzzling, because methylation causes cytosine hypermutability and hence is likely to produce del...

    A genome-wide search for eigenetically regulated genes in zebra finch using MethylCap-seq and RNA-seq
    Sandra Steyaert, Jolien Diddens, Jeroen Galle, Ellen De Meester, Sarah De Keulenaer, Antje Bakker, Nina Sohnius-Wilhelmi, Carolina Frankl-Vilches, Annemie Van der Linden, Wim Van Criekinge, Wim Vanden Berghe & Tim De Meyer
    Learning and memory formation are known to require dynamic CpG (de)methylation and gene expression changes. Here, we aimed at establishing a genome-wide DNA methylation map of the zebra finch genome, a model organism in neuroscience, as well as identifying putatively epigenetically regulated genes. RNA- and MethylCa...

    DNA methylation profiling of primary neuroblastoma tumors using methyl-CpG-binding domain sequencing
    Decock A et al.
    Comprehensive genome-wide DNA methylation studies in neuroblastoma (NB), a childhood tumor that originates from precursor cells of the sympathetic nervous system, are scarce. Recently, we profiled the DNA methylome of 102 well-annotated primary NB tumors by methyl-CpG-binding domain (MBD) sequencing, in order to ide...

    Identification and validation of WISP1 as an epigenetic regulator of metastasis in oral squamous cell carcinoma
    Clausen MJ, Melchers LJ, Mastik MF, Slagter-Menkema L, Groen HJ, van der Laan BF, van Criekinge W, de Meyer T, Denil S, Wisman GB, Roodenburg JL, Schuuring E
    Lymph node (LN) metastasis is the most important prognostic factor in oral squamous cell carcinoma (OSCC) patients. However, in approximately one third of OSCC patients nodal metastases remain undetected, and thus are not adequately treated. Therefore, clinical assessment of LN metastasis needs to be improved. The p...

    DNA methylation in an engineered heart tissue model of cardiac hypertrophy: common signatures and effects of DNA methylation inhibitors
    Stenzig J et al.
    DNA methylation affects transcriptional regulation and constitutes a drug target in cancer biology. In cardiac hypertrophy, DNA methylation may control the fetal gene program. We therefore investigated DNA methylation signatures and their dynamics in an in vitro model of cardiac hypertrophy based on engineered heart...

    ∆ DNMT3B4-del Contributes to Aberrant DNA Methylation Patterns in Lung Tumorigenesis
    Mark Z. Ma, Ruxian Lin, José Carrillo, Manisha Bhutani, Ashutosh Pathak, Hening Ren, Yaokun Li, Jiuzhou Song, Li Mao
    Aberrant DNA methylation is a hallmark of cancer but mechanisms contributing to the abnormality remain elusive. We have previously shown that ∆DNMT3B is the predominantly expressed form of DNMT3B. In this study, we found that most of the lung cancer cell lines tested predominantly expressed DNMT3B isoforms without e...

    Prenatal Exposure to DEHP Affects Spermatogenesis and Sperm DNA Methylation in a Strain-Dependent Manner.
    Prados J, Stenz L, Somm E, Stouder C, Dayer A, Paoloni-Giacobino A
    Di-(2-ethylhexyl)phtalate (DEHP) is a plasticizer with endocrine disrupting properties found ubiquitously in the environment and altering reproduction in rodents. Here we investigated the impact of prenatal exposure to DEHP on spermatogenesis and DNA sperm methylation in two distinct, selected, and sequenced mice st...

    Quality evaluation of methyl binding domain based kits for enrichment DNA-methylation sequencing.
    De Meyer T, Mampaey E, Vlemmix M, Denil S, Trooskens G, Renard JP, De Keulenaer S, Dehan P, Menschaert G, Van Criekinge W
    DNA-methylation is an important epigenetic feature in health and disease. Methylated sequence capturing by Methyl Binding Domain (MBD) based enrichment followed by second-generation sequencing provides the best combination of sensitivity and cost-efficiency for genome-wide DNA-methylation profiling. However, existin...

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