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Parasite culture

JH Johanna Hoshizaki
SA Sophie H. Adjalley
VT Vandana Thathy
KJ Kim Judge
MB Matthew Berriman
AR Adam J. Reid
ML Marcus C. S. Lee
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P. falciparum parasites (3D7 strain) were grown as asexual blood-stage cultures in RPMI media with AlbuMAX® (Gibco) and supplemented with GlutaMax® (Gibco), Gentamicin (Gibco) and HEPES (pH 7) with O+ human erythrocytes at 3% haematocrit. Cultures were maintained at 37 °C in a gaseous environment of 3% CO2, 1% O2 and 96% N2. Parasitemia and stages were monitored using Giemsa staining and microscopy. Parasite samples for long-read RNA-seq were harvested from a mixed-staged Pf3D7 culture. Parasite samples for short-read RNA-seq were harvested from synchronised Pf3D7 cultures at different time points around the intra-erythrocytic development cycle (0, 8, 16, 24, 32, 40 and 48 hours) with four replicates for each time point. RNA was extracted from parasites using Trizol as previously described [52].

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