RNASeq of Plasmodium falciparum 3D7 WT compared to MSP2 KO
RNASeq of Plasmodium falciparum 3D7 WT compared to MSP2 KO
Cultures were tightly synchronised and harvested at late schizont. Cultures were pelleted, resuspended in TriZol (Invitrogen) and incubated for 5 minutes at 37°C before either immediate RNA extraction or freezing at -80°C. Chloroform (Sigma Aldrich) was added, mixed and then spun at 12,000g for 30 minutes at 4°C. Supernatant was collected and mixed with an equal volume of 70% Ethanol and processed using the RNeasy Kit (Qiagen) to extract purified RNA. RNA was DNAse l (Qiagen) treated for 30 mins at room temperature and cleaned up on RNeasy columns according to the manufacturers protocol. Absence of gDNA was checked by qPCR of gDNA and DNAse I treatment repeated if necessary. Library construction and sequencing was performed by Victorian Clinical Genetics Services (VCGS) Australia. Library was prepared using llumina stranded total RNA kit with Ribo -Zero Plus. QC was performed after libary prepration and prior to sequencing.