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The Methylation Ligation-dependent Macroarray (MLM), an array-based analysis, was designed in order to measure methylation levels of 58 genes previously described as putative biomarkers of cancer.
We designed two independent primer panels to measure methylation levels at selected genomic regions.
In array-based methylation experiments, both Beta-value and M-value statistics are used as metrics to measure methylation levels.
To measure methylation levels, bisulfite conversion has been combined with restriction analysis (COBRA [ 5]), base-specific cleavage and mass spectrometry [ 6], real-time PCR (MethyLight, [ 7]), and pyrosequencing [ 8].
In the present study, we first used both 5-mdC and LINE-1 methodologies to measure methylation levels of blood DNA in a subset of 19 breast cancer cases and 18 controls.
In the first experiment, we measured DNA methylation levels at 1,536 CpG sites in cord blood and placenta of 22 individuals using a custom-designed DNA methylation array (which uses the "GoldenGate" assay to measure methylation levels; Illumina, Inc. USA, see Methods and [ 32]).
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Illumina's Infinium Methylation Assay is used to measure methylation level at the single-CpG-site and therefore can offer the highest resolution for identifying the various tissue types and solving many different forensic cases (Table 2).
None of the studied loci showed a correlation between the measured methylation levels and age of the DNA sample.
In phase I, we measured methylation levels at 1,505 CpG sites in treatment-naïve leukocyte DNA from 132 never-smoker PaC patients and 60 never-smoker healthy controls.
Both methods are highly sensitive and capable of measuring methylation levels.
We fit a mixed-effects model to account for repeatedly measured methylation levels.
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measuring methylation levels
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used methylation levels
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