Thymidine kinase as a negative selectable marker in Leishmania major.

We have tested whether the thymidine kinase gene of herpes simplex virus 1 (HSV-1 tk) could function as a negative selectable marker in Leishmania major. Several drug resistance markers have been used for positive selection of transfected DNA in trypanosomatids, including neomycin phosphotransferase (NEO; refs. 1-4) and hygromycin phosphotransferase (HYG; refs. 5,6). Negative selectable markers, that is genes whose expression is lethal to cells, have also proven useful for a variety of purposes in prokaryotic and eukaryotic systems. For example, negative selections have been used to enrich for homologous versus random recombinants in cultured mammalian cells [7], to ablate specific cell lineages in transgenic mice after fusion of the negative marker to a cell type-specific enhancer/promoter [8-11], and to select for muta-