TCGAbiolinks is an R package, which is licensed under the General Public License (GPLv3), and is freely available through the Bioconductor repository (21 (link)). By conforming to the strict guidelines for package submission to Bioconductor, we were able to utilize and incorporate existing R/Bioconductor packages and statistics to assist in identifying differentially altered genomic regions defined by mutation, copy number, expression or DNA methylation; to reproduce previous TCGA marker studies; and to integrate data types both within TCGA and across other data types outside of TCGA. TCGAbiolinks consists of functions that can be grouped into three main levels: Data, Analysis and Visualization. More specifically, the package provides multiple methods for the analysis of individual experimental platforms (e.g. differential expression analysis or identifying differentially methylated regions or copy number alterations) and methods for visualization (e.g. survival plots, volcano plots and starburst plots) to facilitate the development of complete analysis pipelines. In addition, TCGAbiolinks offers in-depth integrative analysis of multiple platforms, such as copy number and expression or expression and DNA methylation, as demonstrated and applied in our recent TCGA study of 1122 gliomas (6 ). These functions can be used independently or in combination to provide the user with fully comprehensible analysis pipelines applied to TCGA data. A schematic overview of the package is presented in Figure 2 . We will describe each of the three main levels (Data, Analysis and Visualization) below, highlighting the importance and utility of each associated function and subfunction. We will then introduce four tumor case studies, which will help clarify the utility of TCGAbiolinks for the reader. We have also compiled an in-depth vignette, which describes every function in detail. Here, we will summarize the main functions.
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DNA Methylation
DNA Methylation
DNA methylation is a fundamental epigenetic process that involves the addition of methyl groups to DNA molecules, typically at cytosine residues.
This modification can influence gene expression, chromatin structure, and genome stability without altering the underlying DNA sequence.
Dysregulation of DNA methylation patterns has been implicated in various disease states, including cancer, neurological disorders, and imprinting disorders.
Understanding the complex patterns and functions of DNA methylation is crucial for advancing biomedical research and developing targeted therapies.
PubCompare.ai offers an AI-driven comparison tool to help researchers streamline their DNA methylation experiments, identify the most effective protocols and products from published literature, preprints, and patents, and optimize their research for better results.
This modification can influence gene expression, chromatin structure, and genome stability without altering the underlying DNA sequence.
Dysregulation of DNA methylation patterns has been implicated in various disease states, including cancer, neurological disorders, and imprinting disorders.
Understanding the complex patterns and functions of DNA methylation is crucial for advancing biomedical research and developing targeted therapies.
PubCompare.ai offers an AI-driven comparison tool to help researchers streamline their DNA methylation experiments, identify the most effective protocols and products from published literature, preprints, and patents, and optimize their research for better results.
Most cited protocols related to «DNA Methylation»
Copy Number Polymorphism
DNA Methylation
Genome
Glioma
Mutation
Neoplasms
The DNA methylation data considered in this work were all generated using Illumina’s Infinium Human Methylation 450 k beadchip. Full details of this technology are described in Bibikova et al. (2011) (link) and Sandoval et al. (2011) (link). Briefly, the methylation value of each probe follows an approximate -valued distribution, with constrained to lie between 0 (unmethylated locus) and 1 (methylated). This follows from the definition of as the ratio of methylated to combined intensity values, i.e
where and are the unmethylated and methylated intensity values of the probe (averaged over bead replicates) and is a small correction term to regularize probes of low total signal intensity (i.e. probes with after background subtraction). Throughout we used non–background-corrected DNAm data. Of the 485 577 probes, 72% are of a type2 design in which the and measurements are obtained in different colour channels, while the rest (28%) of the probes are of the old type1 design in which both and measurements are obtained in the same colour channel. Importantly, type1 and type2 probes differ significantly in terms of CpG density, with CpGs mapping to CpGs islands overrepresented among type1 probes (Bibikova et al., 2011 (link); Sandoval et al., 2011 (link)).
where and are the unmethylated and methylated intensity values of the probe (averaged over bead replicates) and is a small correction term to regularize probes of low total signal intensity (i.e. probes with after background subtraction). Throughout we used non–background-corrected DNAm data. Of the 485 577 probes, 72% are of a type2 design in which the and measurements are obtained in different colour channels, while the rest (28%) of the probes are of the old type1 design in which both and measurements are obtained in the same colour channel. Importantly, type1 and type2 probes differ significantly in terms of CpG density, with CpGs mapping to CpGs islands overrepresented among type1 probes (Bibikova et al., 2011 (link); Sandoval et al., 2011 (link)).
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CpG Islands
cytidylyl-3'-5'-guanosine
DNA Methylation
Homo sapiens
All of the public Illumina DNA data were generated by following the standard protocol of Illumina methylation assays, which quantifies DNA methylation levels by the β value. A detailed description of the pre-processing and data normalization steps is provided in Additional file 2 .
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Biological Assay
DNA Methylation
Methylation
We describe several examples using existing methylation data sets as benchmarks for validating the proposed method, in order to demonstrate its clinical or epidemiological utility. First we describe the validation data set S0 used in all examples. Next we describe a laboratory reconstruction experiment, which validates our fundamental proposition that DNA methylation retains substantial information about cell mixtures. Finally we describe the results of applying our methodology to several different target data sets S1. For the head and neck cancer and ovarian cancer data sets, from which bead chip data were available, a linear mixed effects model with a random intercept for bead chip was used to estimate the corresponding row of B 1. For the remaining data sets, no bead chip data were available; consequently, ordinary least squares was used. 250 bootstrap iterations were used for each example and each of the two bootstrap methods of standard error estimation.
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Cancer of Head and Neck
Cells
DNA Chips
DNA Methylation
Methylation
Ovarian Cancer
Reconstructive Surgical Procedures
500 ng of genomic DNA from each sample was treated with sodium bisulfite in duplicate, using the EZ96 DNA methylation kit (Zymo Research, CA, USA) following the manufacturer’s standard protocol. Genome-wide DNA methylation was assessed using the Illumina Infinium HumanMethylation450 BeadChip (Illumina Inc, CA, USA) according to manufacturer’s instructions. Illumina GenomeStudio software was used to extract the raw signal intensities of each probe (without background correction or normalization).
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DNA Methylation
Genome
sodium bisulfite
Most recents protocols related to «DNA Methylation»
The ACADVL gene encodes for very long-chain acyl-CoA dehydrogenase (VLCAD), which functions within mitochondria and is essential for fatty acid oxidation. HCFs were prepared for western blot analysis of VLCAD, caspase-3 (CASP3), cytochrome c (Cyto C), lamin B1, β-actin, and Arp2 with the internal reference protein glyceraldehyde 3-phosphate dehydrogenase (GAPDH) before knockdown of the ACADVL gene. The above proteins were quantified again after knockdown of ACADVL. Detailed methods of the western blotting and knockdown procedure are provided in Additional file 2 .
Knockdown of DNMT1 leads to generally decreased DNA methylation and activates cascades of genotoxic stress [31 (link)] in cells, resulting in signal transduction unrelated to cardiac fibrosis. Thus, we preferred to downregulate the ACADVL gene expression to simulate the HIIT-associated inhibition of human cardiac fibroblast activities.
Knockdown of DNMT1 leads to generally decreased DNA methylation and activates cascades of genotoxic stress [31 (link)] in cells, resulting in signal transduction unrelated to cardiac fibrosis. Thus, we preferred to downregulate the ACADVL gene expression to simulate the HIIT-associated inhibition of human cardiac fibroblast activities.
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Actin-Related Protein 2
Actins
Acyl-Coa Dehydrogenase Very Long Chain Deficiency
Caspase 3
Cells
Cytochromes c
DNA Methylation
DNMT1 protein, human
Fatty Acids, Essential
Fibroblasts
Fibrosis
Gene Expression
Gene Knockdown Techniques
Genes
Genotoxic Stress
Glyceraldehyde-3-Phosphate Dehydrogenases
Heart
lamin B1
Long-Chain-Acyl-CoA Dehydrogenase
Mitochondria
Proteins
Psychological Inhibition
Signal Transduction
Western Blot
Bisulfite analysis was performed using Bismark-bowtie 2 (https://www.bioinformatics.babraham.ac.uk/projects/bismark/ ). We used the default Bowtie2 implementation of Bismark with the specifications that only uniquely mapped reads should be aligned. All alignments were performed with high stringency allowing for only one base mismatch (n = 1). We also clearly specified that no discordant pairs of the pair-end reads should be aligned. DNA methylation in the CG, CHG, and CHH contexts was calculated by dividing the total number of aligned methylated reads by the total number of methylated plus un-methylated reads.
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DNA Methylation
hydrogen sulfite
Differentially methylated regions were called using the DMRcaller R package v1.22.045 . Given the low level of correlation of DNA methylation observed in P. tricornutum11 (link),27 (link) and sequencing coverage in all three cell lines, only cytosines with coverage > =5X in all three lines were kept for further analysis and the bins strategy was favored over other built-in DMRcaller tools. DMRs were defined as 100 bp regions with at least an average 20% loss/gain of DNA methylation in either one of the DNMT5:KOs compared to the reference strain. The ‘Score test’ method was used to calculate statistical significance and threshold was set at p < 0.01. In addition, to distinguish isolated differentially methylated cytosines from regions with significant loss of DNA methylation, an hypoDMR must contain at least methylated 2 CpG in the reference strain.
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BP 100
Cell Lines
Cytosine
DNA Methylation
Strains
To compare and validate TEEM-Seq against more traditional methods of analysis for DNA methylation, we prepared TEEM-Seq libraries from many of the same starling DNA samples that underwent WGEM-Seq and RRBS. For WGEM-Seq, extra material from three of the EM-seq libraries used for bait capture were confirmed clear of adapter dimers on the TapeStation, pooled and sequenced at 2x150 bp reads with 5% PhiX spike-in in 3/4th of a lane (82.5 Gb) on a HiSeq 4000 at Novogene with the same parameters to provide a whole-genome sequence and partial methylome analysis comparison to our targeted sequencing. For a previously unpublished RRBS study, we used NuGEN (now Tecan) Ovation Methyl-Seq kits to prepare RRBS libraries (with added Promega unmethylated cl857 Sam7 lambda control spike-in of 1 ng per 80–120 ng of sample), which were sequenced in several pools of 16 samples per lane (25 Gb) at 1x100 bp reads with 10% PhiX spike-in on a HiSeq 2500 at the Cornell Institute of Biotechnology Genomics Core, Ithaca, NY. Libraries for RRBS were prepared according to kit protocols. Briefly, samples were digested with MspI at 37°C, ligated with bisulfite-compatible barcoded adaptors at 25°C, end repaired at 60°C, bisulfite converted with the Qiagen EpiTect Fast Bisulfite Conversion Kit, PCR amplified with NuGEN’s standard 12 cycles and 60°C annealing, and purified with Agencourt beads for sequencing. Eight of these samples used previously for RRBS were included in the present study, though one failed enzymatic conversion (mean lambda conversion rate from the original RRBS analysis of these samples was 1.43%). Data from the remaining seven samples (from different sequencing pools) were used as a comparison to the TEEM-Seq and WGEM-Seq derived data generated from the same individual starlings.
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DNA Library
DNA Methylation
Enzymes
Epigenome
Genome
hydrogen sulfite
Promega
Sturnidae
A pool of 48 libraries contained 39 starling samples, two house sparrow (Passer domesticus) samples (to compare bait performance in any conserved regions because we planned to use this bait supplier and protocol with another study on house sparrows), and seven replicate libraries. Eight superb starling samples were used in a previously unpublished DNA methylation study that used RRBS. Three of these eight were included in duplicate libraries and one in triplicate libraries, and the house sparrow samples were also run in duplicate. The remaining 31 additional superb starling samples were included to assess target capture performance when scaling up to a pool of 48 libraries.
We followed the myBaits v4.01 protocol, but lowered the default hybridization and clean up temperature from 65°C to 63°C as myBaits recommends for fragmented DNA. We note, however, that a temperature of 65°C works similarly well with EM-seq converted libraries [24 ]. Hybridization beads bound to the pooled library-blocker mix were cleaned with three washes, and the resuspended bead-bound DNA was then amplified with 16 PCR cycles in a KAPA HiFi reaction using a 60°C annealing temperature and a one minute extension step at 72°C. The amplified capture pool was subsequently cleaned up with AMPure XP beads and sequenced at 2x150 bp reads with 5% PhiX spike-in in one lane (110 Gb) of an Illumina HiSeq 4000 at the Novogene Corporation, Sacramento, CA.
We followed the myBaits v4.01 protocol, but lowered the default hybridization and clean up temperature from 65°C to 63°C as myBaits recommends for fragmented DNA. We note, however, that a temperature of 65°C works similarly well with EM-seq converted libraries [24 ]. Hybridization beads bound to the pooled library-blocker mix were cleaned with three washes, and the resuspended bead-bound DNA was then amplified with 16 PCR cycles in a KAPA HiFi reaction using a 60°C annealing temperature and a one minute extension step at 72°C. The amplified capture pool was subsequently cleaned up with AMPure XP beads and sequenced at 2x150 bp reads with 5% PhiX spike-in in one lane (110 Gb) of an Illumina HiSeq 4000 at the Novogene Corporation, Sacramento, CA.
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Crossbreeding
DNA Library
DNA Methylation
One-Step dentin bonding system
Passeridae
Top products related to «DNA Methylation»
Sourced in United States, Germany, China, Italy, Canada, United Kingdom, Australia, Netherlands
The EZ DNA Methylation-Gold Kit is a product offered by Zymo Research for bisulfite conversion of DNA samples. It is designed to convert unmethylated cytosine residues to uracil, while leaving methylated cytosines unchanged, enabling the detection and analysis of DNA methylation patterns.
Sourced in United States, Germany, United Kingdom, France, China
The EZ DNA Methylation Kit is a product offered by Zymo Research for the bisulfite conversion of DNA. The kit is designed to convert unmethylated cytosine residues to uracil, while leaving methylated cytosines unchanged, enabling the detection of DNA methylation patterns.
Sourced in Germany, United States, France, United Kingdom, Netherlands, Spain, Japan, China, Italy, Canada, Switzerland, Australia, Sweden, India, Belgium, Brazil, Denmark
The QIAamp DNA Mini Kit is a laboratory equipment product designed for the purification of genomic DNA from a variety of sample types. It utilizes a silica-membrane-based technology to efficiently capture and purify DNA, which can then be used for various downstream applications.
Sourced in United States
The Infinium HumanMethylation450 BeadChip is a DNA methylation microarray platform developed by Illumina. It is designed to examine DNA methylation patterns across the human genome at over 450,000 CpG sites.
Sourced in United States, Germany
The EZ DNA Methylation-Direct Kit is a product offered by Zymo Research. It is designed for the conversion of DNA samples for bisulfite sequencing, which is a method used to analyze DNA methylation patterns. The kit provides a rapid and efficient way to convert unmethylated cytosine residues to uracil, while leaving methylated cytosines unchanged, allowing for the detection of DNA methylation status.
Sourced in United States, Germany
The EZ DNA Methylation-Lightning Kit is a tool for converting unmethylated cytosines in DNA samples to uracil, allowing for the detection and analysis of DNA methylation patterns. The kit provides a rapid and efficient method for bisulfite conversion of DNA.
Sourced in United States, Germany
The EZ-96 DNA Methylation Kit is a laboratory product designed for the bisulfite conversion of DNA samples. It facilitates the analysis of DNA methylation patterns through techniques such as DNA sequencing or methylation-specific PCR.
Sourced in Germany, United States, Netherlands, United Kingdom, China, Spain, Italy, Canada, Japan, France, Sweden
The EpiTect Bisulfite Kit is a laboratory equipment product designed for bisulfite conversion of DNA samples. It facilitates the chemical modification of DNA to determine the methylation status of specific genomic regions.
Sourced in United States, Netherlands
The HumanMethylation450 BeadChip is a microarray-based technology used for the analysis of DNA methylation patterns. It provides a comprehensive coverage of CpG sites across the genome, allowing for the assessment of DNA methylation levels at over 450,000 specific locations.
Sourced in Germany, United States, United Kingdom, Spain, Canada, Netherlands, Japan, China, France, Australia, Denmark, Switzerland, Italy, Sweden, Belgium, Austria, Hungary
The DNeasy Blood and Tissue Kit is a DNA extraction and purification product designed for the isolation of genomic DNA from a variety of sample types, including blood, tissues, and cultured cells. The kit utilizes a silica-based membrane technology to efficiently capture and purify DNA, providing high-quality samples suitable for use in various downstream applications.
More about "DNA Methylation"
DNA methylation is a fundamental epigenetic process that involves the covalent addition of methyl groups (-CH3) to cytosine residues within DNA molecules.
This chemical modification can influence gene expression, chromatin structure, and genome stability without altering the underlying genetic sequence.
Dysregulation of DNA methylation patterns has been implicated in the pathogenesis of various disease states, including cancer, neurological disorders, and imprinting disorders.
Understanding the complex patterns and functions of DNA methylation is crucial for advancing biomedical research and developing targeted therapies.
Researchers often utilize a variety of techniques and kits to study DNA methylation, such as the EZ DNA Methylation-Gold Kit, EZ DNA Methylation Kit, QIAamp DNA Mini Kit, Infinium HumanMethylation450 BeadChip, EZ DNA Methylation-Direct Kit, EZ DNA Methylation-Lightning Kit, EZ-96 DNA Methylation Kit, EpiTect Bisulfite Kit, and DNeasy Blood and Tissue Kit.
PubCompare.ai offers an AI-driven comparison tool to help researchers streamline their DNA methylation experiments, identify the most effective protocols and products from published literature, preprints, and patents, and optimize their research for better results.
By utilizing this intelligent protocol comparison feature, researchers can enhance the accuracy and efficiency of their DNA methylation studies, leading to more meaningful insights and potential breakthroughs in biomedicine.
This chemical modification can influence gene expression, chromatin structure, and genome stability without altering the underlying genetic sequence.
Dysregulation of DNA methylation patterns has been implicated in the pathogenesis of various disease states, including cancer, neurological disorders, and imprinting disorders.
Understanding the complex patterns and functions of DNA methylation is crucial for advancing biomedical research and developing targeted therapies.
Researchers often utilize a variety of techniques and kits to study DNA methylation, such as the EZ DNA Methylation-Gold Kit, EZ DNA Methylation Kit, QIAamp DNA Mini Kit, Infinium HumanMethylation450 BeadChip, EZ DNA Methylation-Direct Kit, EZ DNA Methylation-Lightning Kit, EZ-96 DNA Methylation Kit, EpiTect Bisulfite Kit, and DNeasy Blood and Tissue Kit.
PubCompare.ai offers an AI-driven comparison tool to help researchers streamline their DNA methylation experiments, identify the most effective protocols and products from published literature, preprints, and patents, and optimize their research for better results.
By utilizing this intelligent protocol comparison feature, researchers can enhance the accuracy and efficiency of their DNA methylation studies, leading to more meaningful insights and potential breakthroughs in biomedicine.