EpiTect Whole Bisulfitome Kit (25)

The QIAGEN EpiTect Whole Bisulfitome Kit is designed for whole genome amplification of bisulfite converted DNA for downstream PCR analyses. The kit employs Multiple Displacement Amplification (MDA) technology, built on proven REPLI-g technology and adapted to the specific requirements of bisulfite converted DNA, which has smaller fragment sizes and altered nucleotide composition due to bisulfite conversion. The amplification reaction proceeds at 28°C for 8 hours in a 40 µl reaction volume, using Epitect bisulfite converted DNA as the starting material with a maximum input volume of 10 µl and a starting amount greater than 50 ng.

A uniquely processive DNA polymerase with 3'→5' exonuclease proofreading activity maintains high fidelity during replication, and the kit provides highly uniform amplification across the entire converted genome with negligible sequence bias. Typical DNA yields are 1–3 µg per reaction, sufficient for 100–300 real-time or end-point PCR analyses from a single EpiTect bisulfite reaction when estimating 10 ng of amplified DNA per reaction. The reaction is set up in a convenient one-step procedure: bisulfite converted template DNA is incubated with a mixture of reaction buffer and REPLI-g DNA polymerase, and after heat inactivation of the polymerase, the amplified DNA is ready for analysis or storage.

Best results are obtained in combination with EpiTect Plus Bisulfite Kits, whose uniquely formulated DNA Protect system enables effective DNA denaturation needed for complete cytosine conversion while preventing the excessive DNA fragmentation associated with high-temperature, low-pH bisulfite conditions. Bisulfite-treated DNA generated with EpiTect Bisulfite Kits is stable for 3 years at -20°C before use in methylation specific PCR. QIAGEN recommends designing amplicons of 80–150 base pairs when analyzing DNA amplified with this kit.