I’ve talked quite a bit about bisulfite conversion and DNA methylation analysis in past posts. Aberrant methylation events have significant impacts in terms of incidence of cancer and development disregulation. Researchers studying DNA methylation are often working with DNA from “difficult” tissues such as formalin-fixed, paraffin embedded tissues, which characteristically yield DNA that is more fragmented than that purified from fresh tissue. Traditional methods for bisulfite conversion involve a long protocol, harsh chemicals, and generally yield highly fragmented DNA. The DNA fragmentation may significantly impact the utility of the converted DNA in downstream applications such as bisulfite-specific PCR or bisulfite sequencing.
An ideal bisulfite conversion system would allow for complete conversion of a DNA sample in a short period of time, provide high yield of DNA, minimally fragment the DNA, work on a wide range of input DNA amounts (from a wide variety of sample types), and, while we’re at it, be easy to use and to store. Whew! That’s quite the list.
Promega now offers a bisulfite conversion system that fits the bill quite nicely. The system is called MethylEdge™ Bisulfite Conversion System and is a quite handy system for delivering high-quality, amplifiable converted DNA for downstream analysis. The system provides all the reagents necessary to perform bisulfite conversion in a column-based format with at least 90% conversion efficiency in less than two hours. The converted DNA is significantly less fragmented than DNA converted with other kits on the market, so you are left with more intact DNA for downstream applications. We’ve tested this kit with DNA purified from FFPE tissue and have found it to work extremely well in preserving the DNA integrity of these samples, too.
The protocol is fairly straightforward. Add ethanol to your wash buffer, dispense your DNA into aliquots (100pg-2µg of input DNA), and prepare your DNA methylation controls. To perform the conversion, simply add conversion reagent to your sample and incubate in a heat block. Cleanup reagents are included, too, and that step is a simple wash over a column. Elute the sample and you’re set. Samples are ready for downstream analysis. Oh, and did I mentioned that the kit can be stored at room temp? How convenient is that?!
Overall, this kit provides a rapid, efficient, complete method for performing bisulfite conversion while maintaining DNA integrity. The DNA controls that are available also help provide a level of quality control both at the conversion step and in post-conversion analysis. If you are routinely performing bisulfite conversion in your lab and are looking for a method that does not degrade your DNA (or that does not require a huge amount of input DNA), this may just be the method for you. Coupled with upstream DNA purification and quantitation reagents and downstream amplification technologies, the MethylEdge™ System gives you the cutting edge in DNA methylation analysis.