Biobanks store valuable clinical samples that could provide a wealth of information about health and disease, but the secrets contained in these samples have been tough to get at with current technologies. But now, NIH researchers report a rapid new method called Tissue Accessible Chromatin (TACh) that could unlock these precious samples for chromatin analyses.
DNaseI is the usual go-to tool for looking at chromatin accessibility, but it requires large amounts of fresh samples and an hour-long prep time to isolate nuclei. And there’s a narrow concentration range that it will work within. Clinical samples, though, are typically available only in minute amounts, often with unknown cell numbers, and are frozen solid. A method called FAIRE can identify regulatory elements in frozen samples, but its success depends on many factors and doesn’t always provide consistent results.
That’s why the researchers developed TACh, a quick way to analyze the chromatin state of frozen clinical specimens. Instead of DNaseI, they use Benzonase or Cyanase, both of which can efficiently digest DNA and RNA under harsh conditions.
In a proof-of-concept study, the team performed TACh on frozen mouse livers. They found that pulverizing the frozen tissue before digestion gave the best signal-to-noise ratio. Both Benzonase and Cyanase gave very similar results to each other and pretty similar results to DNaseI, which was done on nuclei isolated from fresh mouse livers for comparison. All three enzymes recognized most of the highly accessible regions, but the variability came in to play with less accessible regions.
The researchers say that Benzonase and Cyanase identified accessible regions that correlated with low nucleosomal occupancy, euchromatin-specific epigenetic marks, and transcription, making TACh an ideal technique for chiseling away at the chromatin info in frozen clinical samples.
Chill out and read the details at Epigenetics & Chromatin, June 2012