Multi-proxy palaeoecological analyses of lake cores from two sites on northern Vancouver Island reveal previously undocumented non-arboreal environments in the region during the late Pleistocene. Radiocarbon, pollen, sedimentary ancient DNA (sedaDNA), diatom, and grain size analyses indicate that Topknot Lake on the west coast of northern Vancouver Island was not glaciated in the last 18,500 years, extending into the hypothesized regional glacial maximum. A cold herb-shrub coastal tundra existed at the site from ca. 17,500–16,000 cal BP with species including willows (Salix), grasses (Poaceae), sedges (Cyperaceae), heathers (Ericaceae), and sagebrush (Artemisia). SedaDNA analysis also supports the presence of rare non-arboreal taxa at Topknot Lake during this interval including Jacob's-ladder (Polemonium), bistort (Bistorta), and wild berries (Rubus). After ca. 16,000 cal BP and through the terminal Pleistocene, pine (Pinus), alder (Alnus), and ferns formed open forests under cool and dry conditions. At Little Woss Lake in the mountains of north-central Vancouver Island, fir (Abies) stands dominated from ca. 14,200–14,100 cal BP, then were replaced by open pine woodland with alder and ferns from ca. 14,100–12,000 cal BP. SedaDNA corroborates these plant taxa as well as indicating grizzly bear (Ursus arctos horribilis) and Chinook salmon (Oncorhynchus tshawytscha) in and around the basin by ca. 14,100 cal BP. Mixed conifer forests of pine, western hemlock (Tsuga heterophylla), and alder spread into the island's interior ca. 12,000–11,100 cal BP during the Pleistocene-Holocene transition. The records from these two lakes demonstrate the diachronous development of postglacial ecosystems on northern Vancouver Island. Furthermore, these data provide key evidence for environments that could have supported human populations on the northwest coast of North America for several millennia during the terminal Pleistocene.