In vivo construction of linear vectors based on killer plasmids from Kluyveromyces lactis: selection of a nuclear gene results in attachment of telomeres.

Linear vectors based on plasmids pGKL1 and pGKL2 from Kluyveromyces lactis were obtained by in vivo recombination in Saccharomyces cerevisiae and selected for integration of the nuclear LEU2 gene. The linear hybrid molecules obtained had no proteins attached to their 5' ends, as is found for native pGKL plasmids. However, telomere-specific sequences were added to the ends of pGKL1. In contrast to the cytoplasmically localized pGKL plasmids, the newly obtained linear hybrid vectors probably replicate within the nucleus and provide evidence that the nuclear LEU2 gene cannot be expressed in the cytoplasm.