Figure 5.

TbMLP can be deleted in the presence of a regulated ectopic copy.A. Deletion of endogenous TbMLP alleles. Map shows location of HindIII sites and the small black line indicates the probe. The Southern blot of genomic DNA digested with HindIII shows interruption of the two endogenous alleles in the conditional null mutants [lanes 1, 2 and 3 are individual clones, the wild type (WT) and the heterozygote (SKO) are shown for comparison].B. Repression of conditional TbMLP expression leads to downregulation within 24 h. Total RNA was isolated and blotted from wild type (WT), heterozygote (SKO) and two conditional null clones (DKO1 and DKO2), +tet, cells incubated with tetracycline (1 μg ml⁻¹), −tet, cells washed (× 3) and removed from tetracycline for 24 h.C. Growth phenotype of conditional null mutant recapitulates the RNAi phenotype. The broken black line in each case indicates the cumulative growth of the cell line maintained on tetracycline. The solid black line indicates the growth of the same line after removal of tetracycline by washing. A representative experiment is shown for each clone.