Supporting Information Schroeder et al. “Origins and genetic legacies of the Caribbean Taino” SI Text 1. Terminology 1.1 The term “Taino”. The term “Taino” is often used collectively to refer to the indigenous ​ inhabitants of the Caribbean (1), but as Keegan and Hofman (2) have pointed out this assumes a ​ ​ ​ level of homogeneity in the indigenous Caribbean that is unwarranted. At the time of contact, the Caribbean was a complex mosaic of different ethnic groups, which had considerable contact with each other, and the mainland (3, 4). Columbus simply referred to the people he met as “indios” in ​ ​ the mistaken belief that he had arrived in Asia. Later reports distinguish between the “peaceful Arawaks” who lived the Greater Antilles and the Bahamas, where they were known as the Lucayans, and the more “warlike Caribs” who inhabited the Lesser Antilles. A third group known as the “Guanahatabey” (and often mistakenly referred to as “Ciboney”) allegedly lived in Western Cuba where they had been pushed by the advancing Arawaks. The “name game” (2) has ​ continued in more recent times; for example, eminent Caribbean archaeologist Irving Rouse distinguished between the “Classic Taino” who lived in eastern Cuba, Hispaniola, and Puerto Rico and the “Eastern and Western Taino” who lived in the northernmost Lesser Antilles and Jamaica and central Cuba, respectively (1). Bearing this in mind, we use the term “Taino” as a ​ ​ shorthand to refer collectively to the group of Arawakan speakers who inhabited large parts of the Caribbean at the time of European contact and the term “Lucayan Taino” or simply “Lucayan” to refer to the indigenous inhabitants of the Bahamas. 1.2 Native American languages. Comparisons of linguistic and genetic variation can be a powerful tool for reconstructing human population history. In 1987, the American linguist Joseph Greenberg (5) proposed a classification scheme that split all Native American languages into ​ three major groups, which he referred to as Amerind, Na-Dene, and Eskimo-Aleut. Greenberg further suggested that this division represented three separate migrations into the Americas. Interestingly, genetic studies seem to support Greenberg’s three-way classification in that they suggest that modern Native Americans descend from at least three streams of Asian gene flow (6, ​ 7). However, this does not imply that Greenberg’s categories and subdivisions are correct. In fact, ​ it should be noted that while his classification scheme has been widely and uncritically used by human geneticists, it has been widely rejected by historical linguists who study Native American languages. As Bolnick et al. (8) pointed out, Greenberg’s method of “multilateral comparison” ​ was fundamentally flawed and its results are, therefore, utterly unreliable. The danger of using a classification system that has been almost universally rejected by specialists in the field is that it might obscure evolutionary relationships between groups or suggest links that do not exist. Therefore, we opted to break with tradition, and go for the more widely accepted language classifications proposed by Lyle Campbell and others (9–11). However, we note that many of the ​ ​ classifications and proposed language families remain tentative. 2. Archaeological context The archaeological site of Preacher’s Cave is located on the northern part of the island of Eleuthera in the Bahamas, next to Jean’s Bay and directly south of a reef system known as the Devil’s Backbone (Fig. S1). The cave received its moniker from a group of Puritans, known as 1 the Eleutherian Adventurers, who shipwrecked there in 1648 and sought refuge in the cave (12). ​ ​ Archaeological surveys and investigations at Preacher’s Cave revealed multiple phases of occupation dating both to the historic and the prehistoric period. The prehistoric inhabitants of the Bahamas were known as the Lucayans (13). The archipelago is thought to have been first settled ​ ​ around 800-900 AD (13, 14); although some culturally modified material and human remains ​ ​ from the site yielded earlier dates (see Table S1). By 1200 AD, the Lucayans and their customs reflect that of the Taino, a broader nexus of cultural traditions joined by powerful chiefdoms among the Greater Antilles, and from this time period forward, they are often referred to as Lucayan Taino (13, 14). ​ ​ A total of six Lucayan primary burials were discovered within the cave. Like many peoples native to Mesoamerica and the Caribbean, caves were sacred spaces for the veneration of ancestors as well as the origin of the cosmos and humankind (15). Of the six burials, three were ​ ​ well preserved and have been described elsewhere (15). The three burials belonged to two adult ​ ​ males and one female, aged 20-35 years at the time of death (15). Radiocarbon dating of the ​ ​ bones yielded dates between 320 and 1260 cal. AD (15). Two of the burials (burials 1 and 3) ​ ​ contained plaited matting; and the man in burial 3 was buried with a culturally modified triton shell and a cache containing 29 sunrise tellin shells, a nodule of red ochre, and a fish bone sacrifier (15). The cache may have been used for ritual body painting and the triton shell may ​ ​ have been a status symbol. The plaited matting and grave furniture may indicate high status individuals (15). To date, the Lucayan Taino graves from Preacher’s Cave represent the most ​ ​ complete archaeologically documented prehistoric burials in the Bahamas and provide the greatest contribution to the understanding of Lucayan Taino deathways and mortuary practices (15–18). ​ 3. Radiocarbon dating To determine the age of sample PC537, we directly dated the specimen using radiocarbon dating. The currently most reliable method is dating collagen from bone or dentine. However, in tropical environments, collagen is often poorly or not preserved at all (19) and as part of the dentine was ​ used for DNA analysis, the remaining sample was very small. Consequently, we turned to the enamel fraction. While collagen can be purified through various acid and base steps (20), ​ ​ separation of exogenous and endogenous carbon in enamel is more difficult. Dental enamel is carbonate-containing apatite (bioapatite) and the most likely source of contamination is expected to be groundwater carbonates, which are chemically indistinguishable from carbonates originally formed in the bioapatite. Chemical pre-treatment of bioapatite has therefore focused on removing carbonates in labile positions on the crystal surfaces and at grain boundaries (21–24). While the ​ ​ methodology is still far from being a standardised procedure, it is thought to provide a reliable terminus ante quem (21, 25, 26). ​ ​ ​ ​ ​ After DNA sampling, specimen PC537 was cleaned using an air-abrader with aluminium oxide powder at 40 psi and minimum powder flow. The remaining dentine fraction (55 mg) was sampled for radiocarbon dating of collagen with a diamond drill. Subsequently, the enamel fraction was removed from the cementum using a diamond drill (at less than 3000 rpm to avoid heating) in order to obtain a very fine powder. Research has shown that grinding or drilling to fine powder increases the reliability of the enamel dates, due to contaminants at the grain boundaries being made accessible to chemical pre-treatment (21). Dating of the dentine fraction ​ ​ followed standard ORAU procedures for collagen extraction and dating (20). However, to avoid ​ ​ 2 excessive sample loss, we skipped the ultrafiltration step. Finally, the sample was freeze-dried before undergoing combustion and graphitisation. For enamel dating there is currently no standard protocol available, however, several experimental protocols have been applied in the past (21, 25). Due to the very small sample size ​ ​⁠ of 125.63 mg, experimental treatments with acetic acid as frequently used on enamel and bone apatite were deemed too risky: Although showing improvements in dating results, they cause large amount of sample loss requiring a starting sample weight of around 1-2 g (21). As research ​ ​ has shown that enamel dates are, if inaccurate, too young rather than too old – even in radiocarbon depleted environments – it was important to increase the likelihood of sufficient enamel powder surviving chemical pre-treatment for radiocarbon dating at ORAU (> 0.4 mg C) to establish a robust terminus ante quem. At the same time, contamination removal was important ​ ​ to obtain a date as close to the true age as possible. Treatments with HCl are easier to control and have been used previously with good results for the time period in question (26). ​ ​ Procedures as presented in (26) were used as a guideline. The enamel powder was treated ​ with 23 ml of 0.01 M HCl solution for 1h at 4°C, then rinsed three times with Milli-Q water. The timing was reduced from previously published 2h, as experiments have shown that 1h is sufficient to run the reaction to completion (as indicated by a neutral pH after treatment). The enamel sample size unfortunately excluded the use of stronger acid solutions. After chemical pre-treatment, the sample was frozen and subsequently freeze-dried for 48h. CO extraction ​2 followed standard ORAU protocol for shell carbonates (20): The enamel sample was digested in ​ ​ ​ vacuo in a Pyrex® reaction vessel alongside a IAEA-C1 marble standard with phosphoric acid (3 ​ ml, 85%). An in-house gas collection system, a Carlo Erba elemental analyzer and a Sercon stable isotope mass spectrometer were used to recycle the CO2. To optimize the conversion of ​ ​ CO of ‘very small’ samples to graphite, the desiccant magnesium perchlorate was added to the ​2 water trap of the reactor rigs as described by Motuzaite-Matuzeviciute et al. (27). While routine ​ ​ samples at ORAU of 0.8-1.8 mg C do not require this addition, it is critical for sample sizes of <0.8 mg C. The graphite targets are dated using the HVEE AMS system at ORAU (28). For ​ ​ subsequent radiocarbon calibration IntCal13 (29) and OxCal 4.2 (30) were used.