In vitro growth assay of parasite asexual development

To assess the biological relevance of PfDPY19 for the asexual blood stage development of the parasite, the growth of two independent clones of PfDPY19 null mutants was compared with that of the wild-type parental line in three biological replicates. Cultures were synchronized at ring stages by sorbitol synchronization and parasitaemias were determined by flow cytometry (FACSCalibur, BD Biosciences, Franklin Lakes, NJ, USA) using the Syto 11 dye to discriminate between infected and uninfected erythrocytes, as described elsewhere (Urbán et al., 2011). Cultures were adjusted to ~1.5% parasitaemia and 1% haematocrit. After incubation for one complete intraerythrocytic cycle (~48 h), parasitaemias were determined again by flow cytometry. Multiplication rates for each strain were calculated as the ratio between the final and the initial parasitaemia.