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MGMT Pyro Kit (48)
Cat. No. / ID: 970061
MGMT Pyro Kit (48)
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This MGMT Pyro Kit is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease or for patient management.
✓ 24/7 automatic processing of online orders
✓ Knowledgeable and professional Product & Technical Support
✓ Fast and reliable (re)-ordering
Features
- Quantitative methylation detection
- Comprehensive results in real time
- Easy interpretation of complex sequence information
- Sequence context as built-in control of the assay
- Flexible post-run analysis
Product Details
The MGMT Pyro Kit is used for quantitative measurement of methylation of four CpG sites in exon 1 of the human MGMT gene. The kit contains primers and reagents for amplification of the genes, plus buffers, primers, and reagents for quantitative methylation detection in real time using Pyrosequencing technology on the PyroMark Q24 System.
Principle
The MGMT Pyro Kit is used for quantitative measurement of methylation in four CpG sites in exon 1 of the human MGMT gene (genomic sequence on chromosome 10 from 131,265,519 to 131,265,537: CGACGCCCGCAGGTCCTCG) in real time using Pyrosequencing technology on the PyroMark Q24 System. The MGMT (O(6)-methylguanine-DNA methyltransferase) gene encodes for an enzyme that facilitates the repair of alkylated DNA by removing alkyl groups from the O6 position of guanine. Increased methylation of the MGMT gene causes reduced expression of the gene and diminished DNA repair activity. MGMT is methylated to 25–50% in numerous cancers including brain, colon, lung, and breast cancers.
Procedure
Bisulfite converted genomic DNA is amplified by PCR and then sequenced through the defined region (see figure " Illustration of MGMT assay"). Sequences surrounding the defined positions serve as normalization and reference peaks for quantification and quality assessment of the analysis.
After PCR using primers targeting the defined region of exon 1, the amplicons are immobilized on Streptavidin Sepharose High Performance beads. Single-stranded DNA is prepared, and the sequencing primers anneal to the DNA. The samples are then analyzed on the PyroMark Q24 System (see figures " Unmethylated DNA from a human glioblastoma sample", " Methylated DNA from a human glioblastoma sample", and " Highly methylated control DNA") .
See figures
Applications
The MGMT Pyro Kit is used for quantitative measurement of methylation in four CpG sites in exon 1 of the human MGMT gene (genomic sequence on chromosome 10 from 131,265,519 to 131,265,537: CGACGCCCGCAGGTCCTCG).
Supporting data and figures
Unmethylated DNA from a human glioblastoma sample.
Unmethylated DNA from a human glioblastoma sample.
Resources
Safety Data Sheets (2)
Kit Handbooks (1)
FAQ
Can PyroMark Gold reagents be vortexed?