Paleogenomics

Paleogenomics is a field of science based on the reconstruction and analysis of

[1955-], who worked on paleogenomics.

Background

Initially, aDNA sequencing involved cloning small fragments into bacteria, which proceeded with low efficiency due to the oxidative damage the aDNA suffered over millennia.

Sequencing methods

Challenges and techniques

Specific alterations arise from:

• Basis mutational patterns sequence data (C->T mutation)
• Crosslinks
• Cytosine deamination (increased towards read termini)
• Depurination
• Genome fragmentation

Specific patterns and onset of these alterations help scientists to estimate the sample's age.

Formerly, scientists diagnosed post-mortem damages using enzymatic reactions or

in double helix of DNA and abasic site (created by C->T mutation).
A single fragment of aDNA can be sequenced in its full length with HTS. With these data we can create a distribution representing a size decay curve that enables a direct quantitative comparison of fragmentation across specimens through space and environmental conditions. Throughout the decay curve it is possible to obtain the median length of the given fragment of aDNA. This length reflects the fragmentation levels after death, which generally increases with depositional temperature.^[4]

Libraries

Two different libraries can be performed for aDNA sequencing using PCR for genome amplification:

• Double-stranded aDNA library (dsDNA library)
• Single-stranded aDNA library (ssDNA library)

The first one is created using the blunt-end approach. This technique uses two different adaptors: these adaptors bind randomly the fragment and it can then be amplified. The fragment that does not contain both adaptors cannot be amplified causing an error source. To reduce this error,

T/A ligation was introduced: this method consists in inserting the A tailing in DNA sample to facilitate the ligation of T tailed adaptors. In this methods we optimize the amplification of the aDNA.

To obtain ssDNA libraries, DNA is first denatured with heat. The obtained ssDNA is then ligated to two adaptors in order to generate the complementary strand and finally PCR is applied.^[4]

aDNA Enrichment

As aDNA may contain bacterial DNA or other microorganisms, the process requires enrichment. In order to separate endogenous and exogenous fractions, various methods are employed:

• Damaged template enrichment: Used when constructing an ssDNA library because this method targets DNA damage. When Bst polymerase fills the nick, the sample is treated with uracil DNA glycosylase and endonuclease VIII. These compounds attack the abasic site. The undamaged DNA remains attached to streptavidin-coated paramagnetic beads and can be separated from the sample. This method is specific for samples from late Pleistocene Neanderthals.^[5]
• Extension-free target enrichment in solution: this method is based on target-probe hybridization. This method requires DNA denaturation and then inserts overlapping tiled probes along target regions. Then, PCR for DNA amplification is used and finally DNA is linked to a biotinylated adaptor. It's useful for samples from Archaic hominin ancestry.
• Solid-phase target enrichment: in this method
  real-time PCR method are used in parallel with shotgun sequencing
  screening.
• Whole-genome enrichment: used for sequencing the entire genome of single individuals. Whole-genome In-Solution Capture (WISC) is used.[6] This method starts with the preparation of a genome-wide RNA probe library from a species with a genome that is closely related to the target genome in the DNA sample.^[4]

Diversification of present-day non-African populations and anatomically modern humans

By now many studies in different fields have led to the conclusion that present-day non-African population is the result of the diversification in several different

Phenotypic variation in humans

Analysis of

biological adaptation

.

Skin colour

Migration of humans out of

selective pressure

on skin colour trait, favouring lighter skin colour at higher latitudes. The two most important genes involved in skin pigmentation are SLC24A5 and SLC45A2. Nowadays the “light skin” alleles of these genes are fixed in Europe but they reached a relatively high frequency only fairly recently (about 5000 years ago).^[7] Such slow depigmentation process suggests that ancient Europeans could have faced the downsides of low vitamin D production, such as musculoskeletal and cardiovascular conditions. Another hypothesis is that pre-agricultural Europeans could have met their vitamin D requirements through their diet (since meat and fish contain some vitamin D)^[8]

Adaptation to agricultural diet

One of the major examples of adaptation following the switch to agricultural diet is the persistence of production of the lactase enzyme in adulthood. This enzyme is essential to digest lactose present in milk and dietary products and its absence leads to diarrhea following the consumption of these products. Lactase persistence is determined predominantly by a single-base mutation in the MCM6 gene and ancient DNA data show that this mutation became common only within the past 5000 years, thousands of years after the beginning of dairying practices.^[7] Thus, even in the case of lactase-persistence there is a huge time delay between the onset of a new habit and the spread of the adaptive allele and so milk consumption may have been restricted to children or to lactose-reduced products.

Another example of mutation positively selected by the switch to agriculture is the number of AMY1 gene copies. AMY1 encodes for the starch-digesting enzyme

The immune system

The human

selective pressure

on different immune-associated genes. Migrations, for example, exposed humans to new habitats carrying new pathogens or pathogen vectors (e.g. mosquitos). Also the switch to agriculture involved exposition to different pathogens and health conditions, both due to the increased population density and to living close to livestock. However, it is difficult to directly correlate particular ancient genome changes to improved resistance to particular pathogens, giving the vastness and complexity of the human immune system. Besides studying directly changes in the human immune system, it is also possible to study the ancient genomes of pathogens, such as those causing This suggests that in the ancient past plague may had been less virulent compared to more recent Y. pestis outbreaks.

A study of

Plants and animals

Many non-hominin

Challenges and future perspectives

The analysis of ancient genomes of anatomically modern humans has, in recent years, completely revolutionized our way of studying population migrations, transformation and evolution. Nevertheless, much still remains unknown. The first and obvious problem related to this kind of approach, which is going to be partially overcome by the continuous improvement of the ancient DNA extraction techniques, is the difficulty of recovering well preserved ancient genomes, a challenge that is particularly observed in Africa and in Asia, where the temperatures are higher than in other colder regions of the world. Further, Africa is, among all the continents, the one that harbors the most

Bioethics

populations

. In addition, paleogenomic studies have the potential to harm community or individual histories and identities, as well as to reveal denouncing information about their descendants. For these reasons, these kind of studies are still a touchy subject. Paleogenomics studies can have negative consequences mainly because of the discrepancies between articulations of ethical principles and practices. In fact, ancestors’ remains are usually considered legally and scientifically as “artifacts”, rather than “human subjects”, which justifies questionable behaviors and lack of engagement from

communities

. Testing of ancestral remains are therefore used in disputes, claims in treaty, repatriation, or other legal cases. The acknowledgement of the importance and susceptibility of this subject is heading towards ethical commitment and guidance applicable to different contexts, in order to preserve ancestral remains’ dignity and avoid ethical issues.

See also

• Paleogenetics
• Archaeogenetics

References