A total of 63 binary polymorphisms and 10 short tandem repeats (STRs) were genotyped on a sample of 2,344 Y chromosomes from 18 Native American, 28 Asian, and 5 European populations to investigate the origin(s) of Native American paternal lineages. All three of Greenberg's major linguistic divisions (including 342 Amerind speakers, 186 Na-Dene speakers, and 60 Aleut-Eskimo speakers) were represented in our sample of 588 Native Americans. Single-nucleotide polymorphism (SNP) analysis indicated that three major haplogroups, denoted as C, Q, and R, accounted for nearly 96% of Native American Y chromosomes. Haplogroups C and Q were deemed to represent early Native American founding Y chromosome lineages; however, most haplogroup R lineages present in Native Americans most likely came from recent admixture with Europeans. Although different phylogeographic and STR diversity patterns for the two major founding haplogroups previously led to the inference that they were carried from Asia to the Americas separately, the hypothesis of a single migration of a polymorphic founding population better fits our expanded database. Phylogenetic analyses of STR variation within haplogroups C and Q traced both lineages to a probable ancestral homeland in the vicinity of the Altai Mountains in Southwest Siberia. Divergence dates between the Altai plus North Asians versus the Native American population system ranged from 10,100 to 17,200 years for all lineages, precluding a very early entry into the Americas.