Project description:The recent development of a semiconductor-based, non-optical DNA sequencing technology promises scalable, low-cost and rapid sequence data production. The technology has previously been applied mainly to genomic sequencing and targeted re-sequencing. Here, we demonstrate the utility of Ion Torrent semiconductor-based sequencing for sensitive, efficient and rapid chromatin immunoprecipitation followed by sequencing (ChIP-seq) through the application of sample preparation methods that are optimized for ChIP-seq on the Ion Torrent platform. We leverage this method for epigenetic profiling of tumor tissues. Examination of histone modifications in mouse dendentic cells stimulated with LPS, matched melanoma derived cell line, melanoma tumor tissue
Project description:The recent development of a semiconductor-based, non-optical DNA sequencing technology promises scalable, low-cost and rapid sequence data production. The technology has previously been applied mainly to genomic sequencing and targeted re-sequencing. Here, we demonstrate the utility of Ion Torrent semiconductor-based sequencing for sensitive, efficient and rapid chromatin immunoprecipitation followed by sequencing (ChIP-seq) through the application of sample preparation methods that are optimized for ChIP-seq on the Ion Torrent platform. We leverage this method for epigenetic profiling of tumor tissues.
Project description:Methylated DNA immunoprecipitation sequencing (MeDIP-Seq) is a widely used approach to study DNA methylation genome-wide. Here, we present a novel MeDIP-Seq protocol compatible with the Ion Torrent semiconductor-based sequencing platform that is scalable and accurately identifies sites of differential DNA methylation. Additionally, we demonstrate that the high-throughput data derived from MeDIP-Seq on the Ion Torrent platform provides adequate coverage of CpG cytosines, the methylation states of which we validated at single-base resolution on the Infinium HumanMethylation450K Beadchip array. We applied this integrative approach to further investigate the role of DNA methylation in alternative splicing and to profile 5-mC and 5-hmC variants of DNA methylation in normal human brain tissue that we observed localize over distinct genomic regions. These applications of MeDIP-Seq on the Ion Torrent platform have broad utility and add to the current methodologies for profiling genome-wide DNA methylation states in normal and disease conditions. MeDIP-Seq on Ion Torrent Platform in HCT116 and Human Brain
Project description:Methylated DNA immunoprecipitation sequencing (MeDIP-Seq) is a widely used approach to study DNA methylation genome-wide. Here, we present a novel MeDIP-Seq protocol compatible with the Ion Torrent semiconductor-based sequencing platform that is scalable and accurately identifies sites of differential DNA methylation. Additionally, we demonstrate that the high-throughput data derived from MeDIP-Seq on the Ion Torrent platform provides adequate coverage of CpG cytosines, the methylation states of which we validated at single-base resolution on the Infinium HumanMethylation450K Beadchip array. We applied this integrative approach to further investigate the role of DNA methylation in alternative splicing and to profile 5-mC and 5-hmC variants of DNA methylation in normal human brain tissue that we observed localize over distinct genomic regions. These applications of MeDIP-Seq on the Ion Torrent platform have broad utility and add to the current methodologies for profiling genome-wide DNA methylation states in normal and disease conditions.
Project description:To investigate the impact of histone variants and modification on gene regulation, we report high-throughput profiles of six histone markers, H2A.Z, H3K4me2, H3K9me3, H3K27me3, H3K27ac, and H3K36me3, by ChIP-Seq in T-47D breast cancer cells. Libraries were sequenced with the Illumina HiSeq 2000 analyzer for 50 bp paired-end reads and over 20 million uniquely aligned reads were collected for each histone marker. To examine the impact of histone modification on gene expression regulation in T-47D cells.
Project description:It remains a challenge to decipher the complex relationship between DNA methylation, histone modification, and the underlying DNA sequence with limited input material. Here, we developed an efficient, low-input, and low-cost method for simultaneous profiling of genomic binding sites of histone modification and methylation status of the underlying DNA at single-base resolution from the same cells in a single experiment by integrating CUT&Tag with tagmentation-based bisulfite sequencing (CUT&Tag-BS). We demonstrated the validity of our method for both active and repressive histone modifications using 250,000 mouse ESCs. CUT&Tag-BS showed similar enrichment patterns of histone modification to those observed in non-bisulfite-treated control; it further revealed that H3K4me1-marked regions are mostly CpG-poor, lack of methylation concordance, and exhibit prevalent DNA methylation heterogeneity among the cells. We anticipate that CUT&Tag-BS will be widely applied to directly address the genomic relationship between DNA methylation and histone modification, especially in low-input scenario with precious biological samples.
Project description:Here we investigate the possible relationship between the propagation of epigenetic information and the developmental cell proliferation during Xenopus embryogenesis. We systemically inhibited cell proliferation during the G1/S-transition in gastrula embryos and followed their development until the tadpole stage. We quantified by mass spectrometry the abundance of a large set of histone modification states, which reflects the developmental maturation of the embryonic epigenome.