hMeDIP-seq
Hydroxymethylated DNA Immunoprecipitation and Sequencing
hMeDIP-Seq is used to study 5hmC modifications (Xu et al., 2011). It is a slight variation of MeDIP-seq, which is based on the original MeDIP method described by Weber et al (Weber et al., 2005). Although this method is technically almost identical to MeDIP-seq, it is treated as a separate method due to the substantially different biological insight it provides. For a comprehensive insight into epigenetic changes, both 5mC and 5hmC modifications should be measured (Kamdar et al., 2016).
Methylated DNA is isolated from genomic DNA via immunoprecipitation. Anti-5hmC antibodies are incubated with fragmented gDNA and precipitated, followed by DNA purification and preparation of a sequencing library. Deep sequencing provides greater genome coverage, representing the majority of immunoprecipitated hydroxymethylated DNA.
Advantages:
- Covers 5hmC in dense and less dense repeat regions
- Antibody-based selection is independent of sequence and does not enrich for 5mC, due to antibody specificity
Disadvantages:
- Base-pair resolution is lower (~150 bp), as opposed to single-base resolution with other methods
- Antibody specificity and selectivity must be tested to avoid nonspecific interaction
- Biased toward hypermethylated regions
Reagents:
Illumina Library prep and Array Kit Selector
Reviews:
Kamdar S. N., Ho L. T., Kron K. J., et al. Dynamic interplay between locus-specific DNA methylation and hydroxymethylation regulates distinct biological pathways in prostate carcinogenesis. Clin Epigenetics. 2016;8:32
Devall M., Roubroeks J., Mill J., Weedon M. and Lunnon K. Epigenetic regulation of mitochondrial function in neurodegenerative disease: New insights from advances in genomic technologies. Neurosci Lett. 2016;625:47-55
Shull A. Y., Noonepalle S. K., Lee E. J., Choi J. H. and Shi H. Sequencing the cancer methylome. Methods Mol Biol. 2015;1238:627-651
References:
Chowdhury B., Seetharam A., Wang Z., Liu Y., Lossie A. C., et al. A Study of Alterations in DNA Epigenetic Modifications (5mC and 5hmC) and Gene Expression Influenced by Simulated Microgravity in Human Lymphoblastoid Cells. PLoS One. 2016;11:e0147514
Devall M., Roubroeks J., Mill J., Weedon M. and Lunnon K. Epigenetic regulation of mitochondrial function in neurodegenerative disease: New insights from advances in genomic technologies. Neurosci Lett. 2016;625:47-55
Kamdar S. N., Ho L. T., Kron K. J., Isserlin R., van der Kwast T., et al. Dynamic interplay between locus-specific DNA methylation and hydroxymethylation regulates distinct biological pathways in prostate carcinogenesis. Clin Epigenetics. 2016;8:32
Bogdanovic O., Smits A. H., de la Calle Mustienes E., Tena J. J., Ford E., et al. Active DNA demethylation at enhancers during the vertebrate phylotypic period. Nat Genet. 2016;48:417-426
Uribe-Lewis S., Stark R., Carroll T., et al. 5-hydroxymethylcytosine marks promoters in colon that resist DNA hypermethylation in cancer. Genome Biol. 2015;16:69
Zhu L., Lv R., Kong L., Cheng H., Lan F. and Li X. Genome-Wide Mapping of 5mC and 5hmC Identified Differentially Modified Genomic Regions in Late-Onset Severe Preeclampsia: A Pilot Study. PLoS One. 2015;10:e0134119
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History: hMeDIP-seq
Revision by sbrumpton on 2017-06-21 07:50:20 - Show/Hide
Hydroxymethylated DNA Immunoprecipitation and Sequencing
hMeDIP-Seq is used to study 5hmC modifications (Xu et al., 2011). It is a slight variation of MeDIP-seq, which is based on the original MeDIP method described by Weber et al (Weber et al., 2005). Although this method is technically almost identical to MeDIP-seq, it is treated as a separate method due to the substantially different biological insight it provides. For a comprehensive insight into epigenetic changes, both 5mC and 5hmC modifications should be measured (Kamdar et al., 2016).
Methylated DNA is isolated from genomic DNA via immunoprecipitation. Anti-5hmC antibodies are incubated with fragmented gDNA and precipitated, followed by DNA purification and preparation of a sequencing library. Deep sequencing provides greater genome coverage, representing the majority of immunoprecipitated hydroxymethylated DNA.
Advantages:- Covers 5hmC in dense and less dense repeat regions
- Antibody-based selection is independent of sequence and does not enrich for 5mC, due to antibody specificity
Disadvantages:- Base-pair resolution is lower (~150 bp), as opposed to single-base resolution with other methods
- Antibody specificity and selectivity must be tested to avoid nonspecific interaction
- Biased toward hypermethylated regions
Reagents:Illumina Library prep and Array Kit SelectorReviews:Kamdar S. N., Ho L. T., Kron K. J., et al. Dynamic interplay between locus-specific DNA methylation and hydroxymethylation regulates distinct biological pathways in prostate carcinogenesis. Clin Epigenetics. 2016;8:32Devall M., Roubroeks J., Mill J., Weedon M. and Lunnon K. Epigenetic regulation of mitochondrial function in neurodegenerative disease: New insights from advances in genomic technologies. Neurosci Lett. 2016;625:47-55Shull A. Y., Noonepalle S. K., Lee E. J., Choi J. H. and Shi H. Sequencing the cancer methylome. Methods Mol Biol. 2015;1238:627-651References:Chowdhury B., Seetharam A., Wang Z., Liu Y., Lossie A. C., et al. A Study of Alterations in DNA Epigenetic Modifications (5mC and 5hmC) and Gene Expression Influenced by Simulated Microgravity in Human Lymphoblastoid Cells. PLoS One. 2016;11:e0147514Devall M., Roubroeks J., Mill J., Weedon M. and Lunnon K. Epigenetic regulation of mitochondrial function in neurodegenerative disease: New insights from advances in genomic technologies. Neurosci Lett. 2016;625:47-55Kamdar S. N., Ho L. T., Kron K. J., Isserlin R., van der Kwast T., et al. Dynamic interplay between locus-specific DNA methylation and hydroxymethylation regulates distinct biological pathways in prostate carcinogenesis. Clin Epigenetics. 2016;8:32Bogdanovic O., Smits A. H., de la Calle Mustienes E., Tena J. J., Ford E., et al. Active DNA demethylation at enhancers during the vertebrate phylotypic period. Nat Genet. 2016;48:417-426Uribe-Lewis S., Stark R., Carroll T., et al. 5-hydroxymethylcytosine marks promoters in colon that resist DNA hypermethylation in cancer. Genome Biol. 2015;16:69Zhu L., Lv R., Kong L., Cheng H., Lan F. and Li X. Genome-Wide Mapping of 5mC and 5hmC Identified Differentially Modified Genomic Regions in Late-Onset Severe Preeclampsia: A Pilot Study. PLoS One. 2015;10:e0134119