FIGURE 2.

Following Plasmodium homologue of the Mini-Chromosome Maintenance Complex Binding Protein (PfMCMBP) knockdown parasite growth was prolonged three hours and parasite proliferation failed. (A) Immunoblot of protein lysates from PfMCMBP^3HADDschizont stage parasites (36-48 h.p.i.) cultured in [+]/[−] 250 nM Shld1 and probed with antibodies against the HA epitope or Histone H3 (loading control). Quantification of immunoblot was performed by volumetric measurement of fluorescence intensity with the LiCor Odyssey system (n = 3, mean ±SD). (B) Replication curves of PfMCMBP^3HADDparasites cultured [+]/[−] Shld1 reveal the essentiality of PfMCMBP for parasite growth (n = 3, mean with ±SDerror bars). (C) PfMCMBP^3HADDparasites were tightly synchronised and grown for 1.2 cycles with eight-hour time points for the first 32 hrs, then 3-hr time points until the egress of the minus Shld1 condition was complete. At each time point parasite-stage was assessed by counting 100 infected-RBCs for each biological replicate using Hemacolor stain of thin smears (n = 3 biological replicates, mean with ±SDerror bars). (D) SYBR Green I staining demonstrated no significant difference in total DNA content following RNAse A treatment (n = 3 biological replicates) on C1-treated schizont parasites (arrested prior to parasitophorous vacuole rupture). (E) Representative pictures from [+]/[−] Shld1 PfMCMBP^3HADDparasites at schizont-stage (38-42 h.p.i) after PFA-fixation and probing with rat anti-HA (1:50) and goat anti-rat conjugated with Alexa 488 (1:1000). Images were taken with Nikon E800 using the 100× objective. Scale bar is 1 μm