As DNA methylation keeps popping up in the discussion of more and more diseases, it’s clear that current analysis techniques can only give us part of the big picture. To help focus on other parts of that picture, a new method has been devised that zooms in on the methylation status of single cells.
The Restriction enzyme-based Single-cell Methylation Assay (RMSA) let’s you peek inside the lives of single cells (we mean individual cells, not ones that don’t have a date for Valentine’s Day), and gives a more accurate snapshot of DNA methylation in your sample.
Most techniques for analyzing and detecting DNA methylation require a ton of DNA to work well, which means using many hundreds or thousands of cells, and really only ends up giving you an averaged methylation profile of your sample population. Those average methylation patterns just won’t get the job done if you are looking at the effects of small changes in methylation levels, or find you have a lot of cell-to-cell differences within your samples.
RMSA can help solve these problems by using the individual reaction sites on 48-well slides for a high-throughput workflow. Here’s the upshot on how it works:
- Single cells are placed in the wells of AmpliGrid slides and lysed.
- The DNA undergoes enzymatic shearing
- DNA is then cleaved by methyaltion-sensitive restriction enzymes (MSREs)
- Sample DNA is amplified using PCR conditions optimized specifically for single-cell conditions. The type of amplification products report whether a site was methylated or not in that individual cell.
According to the development team, led by Axel Schumacher at the Centre for Addiction & Mental Health in Toronto, RMSA is a low cost, high throughput way to study single-cell methylation, and might even be adapted for real-time protocols or used together with whole genome arrays.
Get the inside scoop on RMSA at Nucleic Acids Research, January 2011.