Illumina HiSeq 2500 paired end sequencing; RNA-seq analysis of anti-malarial drug chemotherapy of murine experimental Cerebral Malaria (ECM)
RNA-seq analysis of anti-malarial drug chemotherapy of murine experimental Cerebral Malaria (ECM)
Mice were euthanised prior to infection (d0), at the onset of ECM (d6 p.i. RMCBS ≤15), in late-stage ECM (d7 p.i. RMCBS ≤6), and after responding successfully to anti-malarial drug treatment (d7+AC, d10, d14, d30 and d60 p.i. RMCBS >6). Euthanised mice were transcardially perfused with PBS, brains isolated and stored at -80°C until use. Female and male 8-10 week old C57BL/6 mice were purchased from Charles River.  All mice were maintained in individually ventilated cages at the University of Manchester. Cryopreserved Pb ANKA parasites (PMID: 22216235) were thawed and passaged once through C57BL/6 mice before being used to infect experimental animals. Animals were infected via intravenous injection of 1x104 pRBCs. Peripheral parasite burdens of infected mice were followed from day 3 p.i. by microscopic examination of giemsa stained thin blood smears. The development of, and subsequent recovery from, experimental cerebral malaria (ECM) was assessed using the rapid murine coma and behaviour scale (RMCBS) [PMID: 27091974]. Mice that scored ≤15 on the RMCBS (invariably d6 p.i.) received up to 6 daily intraperitoneal (i.p.) injections of 30mg/kg artesunate (Sigma) and 30mg/kg chloroquine (Sigma) in PBS or, alternatively, PBS alone. Mice were euthanised prior to infection (d0), at the onset of ECM (d6 p.i. RMCBS ≤15), in late-stage ECM (d7 p.i. RMCBS ≤6), and after responding successfully to anti-malarial drug treatment (d7+AC, d10, d14, d30 and d60 p.i. RMCBS >6). Whole brains were homogenised in Trizol, before using lipid tissue RNA easy kits according to the manufacturer's instructions (Qiagen) to isolate RNA. Isolated RNA was treated with DNAse to remove genomic DNA contamination prior to QC analysis and use in RNA-Seq analysis. Truseq stranded mRNA Assay. Paired end.
Experimental Factor: infect
Plasmodium berghei ANKA
Experimental Factor: compound
artesunate and chloroquine (at same dosage)
Experimental Factor: dose
30
Experimental Factor: sampling time point
14