HIGHLIGHTS

• Purified, non-methylated and methylated human DNA (5 µg/20 µl of each) for use as negative and positive control in methylation detection applications.
• Set of control primers designed to amplify non-methylated, methylated, and mixed methylation copies of the death-associated protein kinase 1 gene (DAPK1) following bisulfite conversion.
• Can be used in conjunction with the EZ DNA Methylation™, EZ DNA Methylation-Gold™, and EZ DNA Methylation-Direct™ kits to assess the efficiency of bisulfite-mediated conversion of DNA.

DESCRIPTION

The Human HCT116 DKO Non-methylated DNA is purified from cells that contain genetic knockouts of both DNA methyltransferases DNMT1 (-/-) and DNMT3b (-/-)1. The DNA derived from HCT116 DKO cells has a low level of DNA methylation (< 5%) and can be used as a negative control for DNA methylation analysis (Figure). The Human HCT116 DKO Methylated DNA is purified HCT116 DKO DNA that has been enzymatically methylated at all cytosine positions comprising CG dinucleotides by M.SssI methyltransferase2 (EC 2.1.1.37;) and can be used as a positive control for DNA methylation analysis.

An assay for complete methylation by M.SssI methytransferase. Digestion of non-methylated and methylated HCT116 DKO DNA with restriction enzymes MspI and HpaII. MspI digests both nonmethylated and methylated DNA. HpaII is sensitive to CpG methylation.

Methylated cytosines comprising CG dinucleotides within DNA remain unconverted following bisulfite treatment, whereas non-methylated cytosines are converted to uracil and detected as thymine following PCR. The control primers, DAPK1 primer I and DAPK1 primer II amplify methylated, non-methylated, and mixed methylation copies of the death-associated protein kinase 1 gene and are intended for use after bisulfite conversion of the control DNA. Recovered DNA is ideal for many applications including downstream analyses such as PCR, restriction endonuclease digestion, sequencing, etc.