One of the challenges of working with ancient DNA samples is that damage builds up over time, breaking the double helix structure into smaller and smaller fragments. In the samples we worked with, these fragments are dispersed and mixed with contaminants, making the reconstruction of a genome a major technical challenge.
But a dramatic paper published on Thursday shows that this is not always true. The damage creates progressively smaller fragments of DNA over time. But if they’re trapped in the right kind of material, they’ll stay where they are, essentially retaining some key features of the ancient chromosomes, even as the underlying DNA breaks down. Researchers have now used it to detail the structure of mammoth chromosomes, with some implications for how these mammals regulated several key genes.
DNA meets Hi-C
The backbone of the DNA double helix consists of alternating sugars and phosphates, chemically bonded together (the DNA bases are chemically bonded to these sugars). Damage from things like radiation can break these chemical bonds, with fragmentation increasing over time. When samples reach the age of something like a Neanderthal, very few fragments are longer than 100 base pairs. Since chromosomes are millions of base pairs long, it was thought that this would inevitably destroy their structure, as many of the fragments would simply fall apart.
But this will only be true if the medium they are in allows propagation. And some scientists suspected that permafrost, which preserves the tissues of some now-extinct Arctic animals, could block that spread. So they set out to test this using mammoth tissue, taken from a sample called YakInf that is approximately 50,000 years old.
The challenge is that the molecular techniques we use to probe chromosomes occur in liquid solutions, where the fragments would simply drift away from each other anyway. So the team focused on an approach called Hi-C, which specifically stores information about which pieces of DNA were close to each other. It does this by exposing the chromosomes to a chemical that will bind any pieces of DNA that are in physical proximity. So even if those pieces are fragments, they will stick together until they end up in a liquid solution.
Several enzymes are then used to convert these linked molecules into a single piece of DNA, which is then sequenced. This data, which will contain sequence information from two different parts of the genome, then tells us that those parts were once close to each other within a cell.
Interpretation of Hi-C
By itself, a single piece of data like this is not particularly interesting; two pieces of the genome can randomly end up next to each other. But when you have millions of pieces of data like this, you can start to build a map of how the genome is structured.
There are two basic rules that govern the pattern of interactions we expect to see. The first is that interactions within a chromosome will be more common than interactions between two chromosomes. And, within a chromosome, parts that are physically closer together in the molecule are more likely to interact than those that are farther apart.
So if you’re looking at a particular segment of, say, chromosome 12, most of the loci that Hi-C will find it interacting with will also be on chromosome 12. And the frequency of interactions will increase as you go to the sequences you are closer and closer to the one you are interested in.
Separately, you can use Hi-C to help rebuild a chromosome even if you start with nothing but fragments. But exceptions to the expected pattern also tell us things about biology. For example, genes that are active tend to be in loops of DNA, with the two ends of the loop held together by proteins; the same applies to inactive genes. Interactions within these loops tend to be more frequent than interactions between them, subtly changing the frequency with which two fragments end up bound together during Hi-C.