Extended Data Fig. 1: Gel electrophoresis of PCR products from laboratory clones and mock clinical samples.

a) Multiplex drug resistance and csp PCR, for a selection of laboratory clones, run on 2% agarose gel. Bands are annotated based on expected sizes for each amplicon. Note variable size of csp due to a deletion in the N-terminal domain in 3D7 and variation in the central repeat region. Mixtures 1 and 2 contained, respectively: 3D7 + KH2 (80:20) and KH2 + 3D7 (80:20). b) Multiplex drug resistance and csp PCR, for mock clinical DBS samples, run on 2% agarose gel. Mock clinical DBS were prepared by combining in vitro cultured P. falciparum RBCs with human whole blood, in ratios expected to produce final parasitaemias of 10%, 1%, 0.1% and 0.01% infected RBCs, with 50 μl blotted onto filter papers to mimic clinical DBS. The proportions of human and parasite DNA per sample were assessed by quantitative PCR (Extended Data Fig. 2). Samples were extracted and assessed in duplicate. Although bands stopped being visible in the 0.01% parasitaemia samples on this gel, nanopore sequence coverage was still adequate for drug resistance genotyping. c) msp1 PCR, for a selection of laboratory clones, run on 1% agarose gel. A single fragment of approximately 5Kb is expected. The same samples were used as template DNA as in gel (A). d) msp1 PCR, for mock clinical DBS samples, run on 1% agarose gel. The same samples were used as template DNA as in gel (B). All template DNA was diluted to 5–10 ng/µl; 4 µl was used as input for the multiplex drug resistance and csp PCR, 2 µl was used as input for the msp1 PCR, both to a final reaction volume of 50ul. 4 µl of each PCR reaction was run on the gel. DBS = Dried Blood Spots. Neg = Negative control (Nuclease Free Water used as template).