EpiTect Methyl II Custom PCR Arrays
For profiling DNA methylation of user-defined gene panels using MethylScreen technology
EpiTect Methyl II Custom PCR Arrays allow you to define your own gene list, tailored to your specific research interest. We then find the nearest predicted CpG islands or other differentially methylated regions closest to those selected target genes to identify the required cataloged real-time PCR assays. Rigorous PCR primer verification and quality controlled manufacturing guarantee reliable assay performance. Build your EpiTect Methyl II Custom PCR Arrays using our genomewide DNA methylation primer assays. EpiTect Methyl II Custom PCR Arrays use MethylScreen technology provided under license from Orion Genomics, LLC.
EpiTect Methyl II Custom PCR Arrays provide high sensitivity (see figure EpiTect Methyl II PCR Assay detects methylation in heterogeneous samples). Results are comparable to methylation analysis using bisulfite sequencing (see figure Comparison with bisulfite Sanger sequencing) and can provide verification for genome-wide methylation analysis studies (see figure EpiTect Methyl II PCR Arrays generate data comparable to that from BeadChip platforms). EpiTect Methyl II Custom qPCR Arrays are highly suited for biomarker discovery studies (see figure Candidate breast cancer DNA methylation biomarkers).
DNA methylation plays an important role in gene expression and it occurs almost exclusively in the context of CpG dinucleotides in the form of a covalent attachment of a methyl residue to the cytosine residue. CpG islands are regions with an elevated GC content and a high frequency of CpG dinucleotides which overlap with the promoter region of 60–70% of all human genes. Hypermethylation of CpG islands at gene promoters is mostly associated with gene silencing.
The EpiTect Methyl II PCR Array system, using MethylScreen technology, relies on the differential cleavage of target sequences by 2 different restriction endonucleases whose activities require either the presence or absence of methylated cytosines in their respective recognition sequences. As real-time PCR quantifies the relative amount of DNA remaining after each enzyme digestion, the methylation status of individual genes and the methylation profile across a gene panel are reliably and easily calculated (see figure Pictorial explanation of results). The use and analysis of both restriction digests as well as their PCR amplification allow the analysis of smaller, more heterogeneous samples.
QIAGEN's proprietary bioinformatics algorithm reliably selects which promoter CpG islands will most likely represent the DNA methylation status of your custom gene list. The rigorous wet-bench PCR primer verification and quality-controlled manufacturing ensure high levels of CpG island-specific assay specificity and amplification efficiency. The performance of the assays on the EpiTect Methyl II Custom PCR Arrays is guaranteed when used with the appropriate RT2 SYBR® Green qPCR Mastermix.
EpiTect Methyl II Custom PCR Arrays allow the simultaneous DNA methylation profiling of a panel of gene promoters. Arrays are available in both 96- and 384-well plate formats to analyze the methylation status of up to 24 or 96 genes related to a specific area of research. Simply select the genes and plate format required (see figure EpiTect Methyl II PCR Array formats) and then contact us.
First, add equal amounts of each genomic DNA sample to components of the EpiTect Methyl II DNA Restriction Kit to set up four different restriction digests: mock (Mo), methyl-sensitive (Ms), methyl-dependent (Md), and double (Msd). After digestion and heat inactivation of the enzymes, mix each digest with the appropriate RT2 SYBR Green qPCR Mastermix, aliquot into the appropriate wells of the EpiTect Methyl II PCR Array, and run the recommended cycling program on a real-time PCR instrument.
Determine the CT values for the characterization of each digest with each gene-specific assay using your instrument’s software. Then, paste the values into the Excel-based data analysis template for your array format to calculate the percentage of methylated DNA.
The EpiTect Methyl II PCR System is an innovative technology and versatile tool for:
Furthermore, EpiTect Methyl II PCR Arrays are also powerful tools for studying regulatory mechanisms behind the gene expression changes observed with RT2 Profiler PCR Arrays and Assays.
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