Genomic DNA underwent bisulfite conversion using the EZ DNA Methylation kit (D5004, Zymo Research), followed by amplification, fragmentation, and hybridization to Infinium EPIC 850k Human DNA Methylation BeadChips (20020530, Illumina) according to manufacturer’s instructions at the University of Southern California Molecular Genomics Core. Bioinformatic analysis was performed in R (v4.2.1). SFT or meningioma DNA methylation data were preprocessed using the minifi pipeline30 (link). In brief, probes were filtered and analyzed using normal-exponential out-of-band background correction, nonlinear dye bias correction, p-value with out-of-band array hybridization masking, and β value calculation (β=methylated/[methylated+unmethylated]). Principal component analysis was performed on the β methylation values from minfi pre-processing pipeline in R. Variable probes were identified from the first three principal components (PCs). PCs greater than 4 contributed to less than 5% of β value variance. The top 2000 probes were selected for analysis by ranking the absolute gene loading score values within PCs and the tumors were projected along the first two PCs.
Infinium epic 850k human dna methylation beadchips
Manufactured by Illumina
The Infinium EPIC 850k Human DNA Methylation BeadChips is a lab equipment product designed to analyze DNA methylation patterns across the human genome. It provides comprehensive coverage of CpG sites and allows for the quantification of DNA methylation levels at a large scale.
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Lab products found in correlation
4 protocols using infinium epic 850k human dna methylation beadchips
1
Genome-wide DNA Methylation Profiling
2
DNA Methylation Analysis of FFPE Samples
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The genomic DNA extracted from FFPE samples was bisulfite converted using the EZ DNA Methylation kit (Zymo Research, Irvine, CA, USA) following the manufacturer's recommended protocol. Bisulfite‐converted DNA was amplified, fragmented, and hybridized to Infinium EPIC 850 k Human DNA Methylation BeadChips (Illumina), following the manufacturer's recommended protocol. The methylation data were preprocessed using the ChAMP package (v.1.28.4) in R Bioconductor (version 3.5.3). Quality control and normalization were performed according to the standard pipeline of ChAMP. The reference methylation data (450k) were obtained from GEO (GSE109381) [33 (link)]. The t‐distributed stochastic neighbor embedding (t‐SNE) analysis was performed using Rtsne package version 0.16, and the 428,230 probes common between the 450k and EPIC arrays were used for analysis. The input for the t‐SNE calculation is Pearson correlation, weighted by variance, and the clustering analyses were performed using the beta values of the top 32,000 most variably methylated probes by standard deviation.
3
Tumor DNA Extraction and Methylation Analysis
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Tumor tissue was selectively scraped from unstained slides or punched from formalin‐fixed, paraffin‐embedded (FFPE) blocks using 2.0 mm disposable biopsy punches (Integra Miltex Instruments) to enrich for the highest tumor content possible. Genomic DNA was extracted from this macrodissected tumor tissue using the QIAamp DNA FFPE Tissue Kit (Qiagen). Genomic DNA was bisulfite converted using the EZ DNA Methylation kit following the manufacturer's recommended protocol (Zymo Research). Bisulfite converted DNA was then amplified, fragmented, and hybridized to Infinium EPIC 850k Human DNA Methylation BeadChips following the manufacturer's recommended protocol (Illumina).
4
Bisulfite Conversion and DNA Methylation Analysis
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Genomic DNA underwent bisulfite conversion using the EZ DNA Methylation kit (Zymo Research, cat# D5004), followed by amplification, fragmentation, and hybridization to Infinium EPIC 850k Human DNA Methylation BeadChips (Illumina, cat# 20020530) according to manufacturer’s instructions at the Molecular Genomics Core at the University of Southern California (Los Angeles, CA). Bioinformatic analysis was performed in R (v3.6.1). Meningioma DNA methylation data were preprocessed using the SeSAMe pipeline (Bioconductor v3.10) as previously described13 (link),54 (link). In brief, probes were filtered and analyzed using normal-exponential out-of-band background correction, nonlinear dye bias correction, p-value with out-of-band array hybridization masking, and β value calculation (β=methylated/[methylated+unmethylated]). Meningioma samples were assigned to Merlin-intact, Immune-enriched, or Hypermitotic DNA methylation groups using a support vector machine classifier, as previously described13 (link).
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