characteristics.text: 2025-04-30
characteristics.characteristic: ENA first public
characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: BAH-GU
characteristics.characteristic: INSDC center name
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS23779581
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-14868:MFA2A
characteristics.characteristic: Submitter Id
characteristics.text: ArrayExpress
characteristics.characteristic: broker name
characteristics.text: mixed Pf2004 and Dd2
characteristics.characteristic: cell_line
characteristics.text: mixed stages
characteristics.characteristic: cell_type
characteristics.text: not collected
characteristics.characteristic: collection date
characteristics.text: Protocols: Two culture adapted P. falciparum strains were used: Pf2004 from West Africa and Dd2 from Southeast Asia (29-31). At 26-28 hours post invasion (hpi), highly synchronised parasite cultures were subjected to either mFA (induction medium) or serum supplemented control medium. After 16 and 22 hours of further incubation, respectively, two time points were collected for analysis and samples from both strains pooled per condition. Before proceeding with sample preparation, blood smears were made to check for the presence of live parasites. Samples were processed in batches on ice to limit cell loss. Serum samples were processed first by centrifuging at 2000 rpm for 5 minutes to remove Hypothermasol. Parasites were then washed in 5 volumes of cold 1xphosphate buffered saline (PBS):0.2% BSA and centrifuged at 2000 rpm for 5 minutes. To lyse RBCs, the supernatant was aspirated, and pellets were gently resuspended in 5 volumes of 0.1% saponin. Parasites were incubated in 0.1% saponin for 2-5 minutes on ice. After saponin lysis, parasites were gently washed twice with cold 1xPBS:0.2% BSA at 2000 rpm for 5 minutes. At this point, parasites were very fragile and prone to lysis; therefore, wide pore pipette tips were used from this point onwards. Parasites were counted by flow cytometry (MACSQuant VYB flow cytometer (Miltenyi Biotec)) and by Neubauer haemocytometer. Whilst counting was being performed, the next batch of mFA samples were prepared.  To calculate cell concentration from haemocytometer counts, the following calculation was performed:  Cell concentration (cell/mL) =  (number of cells counted / (number of squares counted x dilution factor)) x 2.5e5  After cells were counted by both methods, the mean of the two methods was taken and samples were diluted to 2.5e5 cells/mL in cold 1xPBS:0.2% BSA. The two time points and two parasite strains were pooled together per condition (i.e., per growth medium) generating two samples: mFA and serum. Two P. falciparum strain were used to investigate sexual commitment: Pf2004/164tdt (referred to as Pf2004) and Dd2 WT (referred to as Dd2). Pf2004/164tdt was previously transfected with the reporter plasmid 164tdTomato under the control of the gametocyte specific promoter, PF3D7_1016900 (48). Asexual parasites were routinely cultured as previously described (49, 50). Parasites were grown in complete medium composed of RPMI 1640 with L-Glutamine, 25 mM HEPES, 5 mM hypoxanthine and 1 mg/mL gentamycin, supplemented with 10% O-positive human serum (Interstate Blood Bank, Inc). To select for stable maintenance of episomes in Pf2004/164tdt, media was supplemented with 4 nM WR99210 (Jacobus Pharmaceuticals). Parasites were grown at 5% haematocrit, unless stated otherwise, and maintained at 0.5-5% parasitaemia. Parasites were maintained at 37 oC and supplied with 1% O2, 5% CO2 and 94% N2. After cell preparation, approximately 1e6 cells/mL were loaded onto the 10X Chromium Chip which was loaded onto the 10X Chromium machine to recover 10,000 cells per inlet.  Amplification, library preparation and Illumina sequencing was performed following the standard 10X protocol.
characteristics.characteristic: description
characteristics.text: plasmodium parasite stage
characteristics.characteristic: developmental stage
characteristics.text: Plasmodium falciparum malaria
characteristics.characteristic: disease
characteristics.text: mixed Pf2004 and Dd2
characteristics.characteristic: genotype
characteristics.text: not collected
characteristics.characteristic: geographic location (country and/or sea)
characteristics.text: minimum fatty acid medium
characteristics.characteristic: growth condition
characteristics.text: not applicable
characteristics.characteristic: isolate
characteristics.text: Plasmodium falciparum 3D7
characteristics.characteristic: organism
characteristics.text: Plasmodium falciparum 3D7
characteristics.characteristic: organism
characteristics.text: parasite only
characteristics.characteristic: organism part
characteristics.text: Plasmodium falciparum 3D7
characteristics.characteristic: scientific_name
characteristics.text: MFA2A
characteristics.characteristic: title