Plasmodium falciparum Sonication ChIP-Seq erythrocytic cycle 0h
Parasites were extracted from erythrocytes by saponin-lysis and crosslinked for 10 min at RT using 1% formaldehyde. Nuclei were isolated and chromatin was fragmented by sonication for a total of 25 cycles of (30s ON and 30s OFF) at high intensity using a Bioruptor UCD-200 (Diagenode, Denville, NJ). Anti-histone H3 (Abcam ab1791) immunoprecipitations were performed using the Chromatin Immunoprecipitation Assay Kit (Millipore, Billerica, MA) according to the manufacturer's protocol. Samples were decrosslinked by O/N incubation at 45°C, followed by RNase A treatment (30 min at 37°C), proteinase K treatment (2 h at 45°C) and DNA purification. Sequence libraries were prepared using the Encore Multiplexing System (NuGen) according to the manufacturer’s instructions, with the following modifications for the high AT-content of the P. falciparum genome: the libraries were amplified for a total of 15 PCR cycles (5 cycles of [15s at 98°C, 30s at 55°C, 30s at 62°C] followed by 10 cycles of [15s at 98°C, 30s at 63°C, 30s at 72°C]) using KAPA HiFi HotStart Ready Mix (Kapa Biosystems, Woburn, MA).