Researchers trace advances in ancient DNA technology


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Credit: IVPP

Over the past 10 years, researchers led by FU Qiaomei from the Institute of Vertebrate Paleontology and Paleoanthropology (IVPP) of the Chinese Academy of Sciences (CAS) have used ancient DNA (aDNA) technology to dig up the history of ancient human populations, especially those in East Asia.

As part of their efforts, the researchers reconstructed the entire genome of two extinct groups of archaic humans: Neanderthals and Denisovans; mapped the history of global population migrations and interactions; discovered the genetic structure of the oldest East Asians; revealed adaptive genetic changes in East Asian Ice Age populations; and traced the formation of population patterns in northern and southern China as well as the origin of the Austronesian population in southern China.

Recently, the FU team reviewed the history of aDNA technology development, discussed current bottlenecks and technical solutions, and assessed the future of the technology.

The study was published in Cell July 21.

A key technological development discussed in the study is high-throughput sequencing, which is a technique for rapidly sequencing large amounts of DNA. It can theoretically sequence all DNA molecules in a sample.

Before high-throughput sequencing became commonplace, the aDNA field relied on polymerase chain reaction (PCR) techniques to sequence a few specific DNA fragments. Researchers could only extract a very limited amount of DNA information with this technology and had difficulty distinguishing genuine aDNA from contaminating DNA.

Complementing advances in sequencing, aDNA researchers have also developed improved methods of constructing DNA libraries to better reflect the characteristics of aDNA. Among these methods, partial uracil-DNA glycosylase (UDG) treatments and the construction of single-stranded DNA libraries are two of the most important. The partial UDG treatment not only preserves some of the DNA damage signal at the ends of the DNA fragments, but also removes most of the aDNA damage in the rest of the molecule. This method improves the accuracy of aDNA sequencing results while preserving the characteristics of aDNA required for validation. The construction of single-stranded DNA libraries allows the direct sequencing of damaged and denatured DNA fragments that may be lost in typical modern DNA library construction techniques.

Advances in library construction have limited effectiveness, however, because aDNA samples often contain a large amount of environmental DNA. Accordingly, useful endogenous aDNA sequences often represent less than 1% of the resulting sequences. To solve this problem, the researchers applied DNA capture technology to the aDNA field by creating DNA and RNA probes with sequences similar to their targets. After adding the probes to the sample extracts, the target aDNA binds to the probes and is then “drafted” from the massive amount of environmental DNA. This technology is widely used in ancient human genome research. Currently, more than two-thirds of ancient human genome data comes from data captured using the “1240k” probe set.

DNA capture technology not only dramatically improves the efficiency of aDNA sequencing; it also makes it possible to recover usable data from samples that would otherwise be too degraded to be analyzed.

More recently, aDNA researchers have pushed the boundaries even further by extracting aDNA directly from “soil” (i.e. sediment). This technology has been applied to samples from the Denisova and Baishiya caves, recovering DNA from ancient humans who lived tens of thousands of years ago.

Despite its successful results, however, the study of aDNA has always been very difficult. aDNA itself is very sensitive to contamination and experiments involving aDNA are extremely delicate. In the past, aDNA extraction and library construction depended almost entirely on manual operations. Recently, a few laboratories around the world have started integrating certain aDNA methods with fully automated robotic pipetting platforms. However, at present, sample pre-processing still requires manual steps. How to integrate this time-consuming and laborious work into an automated system is the next challenge for experimental aDNA technology.

The application of aDNA technology goes far beyond the ancient human genome, of course. Paleomolecular research also covers important topics such as tracing ancient epidemics and symbiotic microbial evolution through ancient microbial information; using ancient epigenetic information to explore the interaction between ancient animals and the environment; and the use of ancient proteins to explore human evolution over long periods of time, including how aDNA influences the physiology and fitness of modern humans.

aDNA is time-stamped genetic information that records the evolution and adaptation of human beings over tens of thousands of years. We now know from aDNA research that several important functional genetic haplotypes derive from archaic human populations. These genes are involved in innate immunity, lipid metabolism, high altitude survivability, and skin color. However, the functions of most of the genetic variants identified by aDNA studies have not yet been determined.

In the future, scientists could use the latest gene-editing technology to build aDNA animal models that would reveal the function of many unknown aDNA variants. This will help us better understand how modern human physiology and health have been affected by the genetic inheritance of our ancient ancestors.

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