In our recent manuscript (https://www.biorxiv.org/content/10.1101/557082v1) we revisit promoter bivalency in naive mouse embryonic stem cells using multiplexed quantitative ChIP.
Why doing multiplexed quantitative ChIP? Multiplexing speaks for itself – many experiments all in one tube. But let’s talk about the need for quantitative ChIP. Here is a boat with an observer. A volcano island rises high above sea level.
Here is the boat again. For the observer, the peak now appears pretty small.
But we as an outside observer can see that it is not the volcano height that changed, it is the sea level that rose up to the peak.
Traditional ChIP normalisation assumes a constant background both on the technical and the biological level. Just like the observer in the boat who does not know the change in sea level, the method is blind to global alterations in histone modification levels. In our study, we find that our multiplexed ChIP operates on a ultra-low technical background and allows us to quantify the true distribution of histone H3K27me3 above the technical background. Strikingly, naive mouse ESC have twice as much H3K27me3 methylated nucleosomes as ESC in Serum, and the gained H3K27me3 modifications distributes broadly across the genome while traditional Polycomb targets, such as bivalent domains retain approximately equal levels.
Read the fill story at https://www.biorxiv.org/content/10.1101/557082v1