We assembled genome-wide data from 271 ancient Iberians, of whom 176 are from the largely unsampled period after 2000 BCE, thereby providing a high-resolution time transect of the Iberian Peninsula. We document high genetic substructure between northwestern and southeastern hunter-gatherers before the spread of farming. We reveal sporadic contacts between Iberia and North Africa by~2500 BCE and, by~2000 BCE, the replacement of 40% of Iberia's ancestry and nearly 100% of its Y-chromosomes by people with Steppe ancestry. We show that, in the Iron Age, Steppe ancestry had spread not only into Indo-European-speaking regions but also into non-Indo-European-speaking ones, and we reveal that present-day Basques are best described as a typical Iron Age population without the admixture events that later affected the rest of Iberia. Additionally, we document how, beginning at least in the Roman period, the ancestry of the peninsula was transformed by gene flow from North Africa and the eastern Mediterranean.

We generated genome-wide ancient DNA data from the Balearic Islands, Sicily, and Sardinia, increasing the number of individuals with reported data from 5 to 66. The oldest individual from the Balearic Islands (~2400 BCE) carried ancestry from Steppe pastoralists that likely derived from west-to-east migration from Iberia, while two later Balearic individuals had less. In Sicily, Steppe pastoralist ancestry arrived by ~2200 BCE in part from Iberia; Iranian-related ancestry arrived by the mid-second millennium BCE contemporary to its previously documented spread to the Aegean; and there was large-scale population replacement following the Bronze Age. In Sardinia, nearly all ancestry derived from the island’s early farmers until the first millennium BCE, with an exception of a third millennium BCE outlier who had primarily North African ancestry and who along with an approximately contemporary Iberian documents widespread Africa-to-Europe gene flow in the Chalcolithic. Major immigration into Sardinia began in the first millennium BCE and today no more than 56–62% of Sardinian ancestry is from its first farmers, which is lower than previous estimates highlighting how Sardinia—like every other region in Europe—has been a stage for major movement and mixtures of people.

The arrival of farming in Europe around 8,500 years ago necessitated adaptation to new environments, pathogens, diets, and social organizations. While indirect evidence of adaptation can be detected in patterns of genetic variation in present-day people, ancient DNA makes it possible to witness selection directly by analyzing samples from populations before, during and after adaptation events. Here we report the first genome-wide scan for selection using ancient DNA, capitalizing on the largest genome-wide dataset yet assembled: 230 West Eurasians dating to between 6500 and 1000 BCE, including 163 with newly reported data. The new samples include the first genome-wide data from the Anatolian Neolithic culture, who we show were members of the population that was the source of Europe's first farmers, and whose genetic material we extracted by focusing on the DNA-rich petrous bone. We identify genome-wide significant signatures of selection at loci associated with diet, pigmentation and immunity, and two independent episodes of selection on height.