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Monday, October 28, 2013

Ancient DNA from prehistoric Bulgaria and Denmark

A paper at the AJHG describes a new cost effective method of significantly increasing the amount of authentic DNA output from ancient samples:

By using biotinylated RNA baits transcribed from genomic DNA libraries, we are able to capture DNA fragments from across the human genome. We demonstrate this method on libraries created from four Iron Age and Bronze Age human teeth from Bulgaria, as well as bone samples from seven Peruvian mummies and a Bronze Age hair sample from Denmark. Prior to capture, shotgun sequencing of these libraries yielded an average of 1.2% of reads mapping to the human genome (including duplicates). After capture, this fraction increased substantially, with up to 59% of reads mapped to human and enrichment ranging from 6- to 159-fold.

This is particularly good news for studies which aim to extract autosomal DNA from hundreds of ancient remains, like Gothenburg University's The Rise project, which I excitedly blogged about earlier this year (see here). In fact, I suspect the aforementioned Danish hair sample is one of the samples from The Rise dataset. The reason I say that is because Morten Allentoft is a co-author on this paper, and he's also doing the DNA analysis for The Rise (see here).

In any case, below are two global Principal Component Analyses (PCA) featuring one of the ancient Bulgarians (V2) and the ancient Dane (M4). The principal components (PC1 & 2) were computed using only modern samples, and then the ancient samples projected onto the PCA space.

The ancient Bulgarian is sitting more or less where modern Bulgarians are usually found on such global plots. On the other hand, the ancient Dane is clearly shifted towards East Asia and the Americas, and as a result clusters with Finns, which I suppose is somewhat unexpected because that never happens with modern Danes. So either there's a problem with the analysis, like, say, projection bias (see below for more details), or this Bronze Age Dane was in fact more eastern in terms of global genetic affinities than modern Danes. The latter might well be true if, for instance, he was a recent descendant of migrants from the east (like present-day Russia), and/or he harbored more Mesolithic hunter-gatherer ancestry than Danes do today.

Now, here are a couple of PCA limited to European samples from the supplemental data PDF, including another ancient Bulgarian (K8) and the same ancient Dane (M4). Unfortunately, PC1 appears to be mostly a reflection of the well documented and very recent founder effect and strong genetic drift experienced by the Finnish population. In other words, it's not saying much more than the fact that the ancient samples weren't affected by the same demographic events and genetic drift as Finns during the past few hundred years. It might have been possible to get more informative results by reducing the Finnish sample to only a handful of the least drifted (ie. least Finnish-like) individuals.

Moreover, it's curious that both ancient samples land in more or less the middle of their respective plots in PC2, despite the fact that they come from very different parts of Europe. I suspect that in these instances projection bias is indeed the problem.

Projection bias is similar to the "calculator effect" (see here), but it affects PCA, especially PCA that include only closely related populations, like from Europe. For more background see Haasl et al. 2012 and Lee et al. 2012.

It's also interesting to note that two of the Iron Age Bulgarians are reported as belonging to mtDNA haplogroups U3b and HV, respectively. Both of these haplogroups are generally accepted to be of Near Eastern origin. They're rare in Europe today (usually <2%), but relatively more common in Bulgaria than most other European countries. This suggests some genetic continuity in Bulgaria from at least the Iron Age to the present. Indeed, U3 has been reported from early Neolithic samples from Germany and Ukraine, which means that the ancient Bulgarian U3 lineage need not have arrived in Europe from the Near East during the metal ages.


Carpenter et al., Pulling out the 1%: Whole-Genome Capture for the Targeted Enrichment of Ancient DNA Sequencing Libraries, The American Journal of Human Genetics (2013),

See also...

More info on two Thracian genomes from Iron Age Bulgaria + a complaint

PCA projection bias in ancient DNA studies


Fanty said...

"This suggests some genetic continuity in Bulgaria from at least the Iron Age to the present:"

Wich contradicts another iron age bulgarian that was checked and who came back as "Sardinian like", at least far different from modern ones.

Davidski said...

It depends how liberal they were with the use of that comparison to Sardinians. Maybe they just really meant Southern European-like? That was probably one of the samples from this study, and my guess is that it was P192-1, who clusters with the Tuscans on the plots in the supplement PDF. That's also the individual who belongs to U3b.

Interestingly, the other two ancient Bulgarians don't cluster with the Tuscans. So maybe there were some major genetic substructures in the Balkans at the time?

Onur said...

Modern Bulgarians are still genetically like Southern Europeans, though most like northern Southern Europeans rather than southern Southern Europeans.

barakobama said...

THe Ancient Thracians and Dacians were known for red hair according to Greeks which is surpassing. There are over 2,000 year old references to their red hair,multiple paintings, and inscriptions of rufus on graves. But modern Bulgarians are almost only dark haired like Greeks. Dominte Y DNa I2a1b, R1b l23 and E1b1b V13 in Bulgarians definitely is not evidence at least on the paternal line of coming to Bulgaria after the iron age and that their paternal lineages are mainly at least from the Neolithic age. The Thracians and Dacian's must be modern Bulgarians main ancestors hopefully they can get more DNA info from Thracians and Dacians to proof this. Red hair only gets up to 1% in R1b1a2a1a L11 dominates Germanic and Italo Celtic west Europe and in Uralic Volga Russia. Maybe Bulgaria is under sampled there has to be at least some red hair. I have seen myself multiple redheads from southeast Europe Serbia, Bosnia, and Romania(just north of Bulgaria).

barakobama said...

"Modern Bulgarians are still genetically like Southern Europeans, though most like northern Southern Europeans rather than southern Southern Europeans."

That's basically what it seems Autosomal DNA says. But you cant generalize southern Europeans because in very ancient times like bronze, copper, and Neolithic age it would be hard for them to be connected and mix. In K7b and globe13 Iberia, Italy, Balkans, and kind of France have higher Meditreaen than the rest of Europe in K12b they have higher Atlantic Med. Iberia has a bit more though than Italy and the Balkans and Sardinia is incredibly high. But Italy and the Balkans also have high amount of Near eastern ancestry while Iberia is more similar to central Europe, British isles, and eastern Europe. There seems to be a huge amount of Near eastern ancestry in Italy and Balkans. In globe13 west Asian and southwest Asian combined for Italy and Greece is about 40%+ and over 30% in the rest of the Balkans. It is centered in Greece, southern Italy and Sicily. It decrease as you go north in Italy and north from Greece and out decrease like crazy from Italy-south central Europe, and Balkans-Hungary and Ukraine.

In globe13, K7b, and K12b in Bulgaria there is much more North Euro and Atlantic Baltic than in Greece and Italy, Around the same as Iberia. Then it rises like crazy once you go a little north from Romania into Ukraine North Euro rises from 40-60% for globe13.

I have noticed huge borders in Europe for percentages in globe13, K7b, and K12b. There are globe13, K7b, and K12b results of pre historic European farmers and hunter gathers. Farmers: Gok4: mtDNA H, Neolithic Funnel Beaker culture, South Sweden, 5,500-4,500 years old, Otzie Y DNA G2a2 L91, mtDNA K1f,Alps Italy, 5,300ybp, early copper age farmer.

Hunter gathers La Brana, mtDNA U5b2c1, Northwest Spain, 7,000ybp, from Mesolithic age, AJV52, 4,000-4,800ybp, Gotland Sweden, Pitted ware culture from Neolithic age, AJV70, mtDNA U4, 4,000-4,800ybp, Gotland, Sweden, Pitted ware culture from Neolithic age.

The Farmers showed results nearly identical to modern Sardina people and overall closest to southern Europeans. They had majority Atlantic Med in K12b, Med in globe13 and K12b. The hunter gathers did not have any extremely close matches in Europe but overall were closest to northern Europeans mainly Finnish and Baltic. In globe13 and K12b they had vast majority North Euro and in K7b Atlantic Baltic was almost 100% for each.

Balkans, Iberia, and Italy(especially Sardinia) may have just the most ancestry from Neolithic farmers in Europe that's all. And later migrations in copper and bronze age probably connected with spread of Indo European languages changed percentages in Europe. It may have raised North Euro and Atlantic Baltic in Europe and also raised and changed percentages of west Asian and southwest Asian. The very high amount of southwest Asian and west Asian in Italy and Balkans is definitely from after Neolithic and probably rom the same source.

Fanty said...

Ancient claims about "red hair" means nothing.

Tacitus claims that all the Germanic people would have in common: massive huge bodies, only usefull for used as a weapon, while its not usefull for hard work. Fierce blue eyes and RED HAIR. For there not beeing much variance in the apearances of the people he claims they must be of pure, unmixed blood blabla.

For Germanic people beeing redhaired he again claims for Picts: Huge massive bodies with red hair, undoubtly the Picts originate in Germania.

Maybe Tacitus needed some glasses. Germanic people have only like 4% red haired people. Its celtic people who reach like 10%. Complete red haired populations like described by Tacitus must be nonsense.

And then there is the first ever description of Slavs: Brutish people with blue eyes and RED HAIR.

Uh really? Almost apears as thats the standart for any kind of Barbarian in the eyes of Romans? ;-)

Years ago I read in a book about Germanic people, an explanation for the "red hair". It claimed the Geranic people used some acid from Oakwood wich would result in the hair getting a red color, wich they suposedly used as a war-paint to look scary....