Sequencing of Antimalarial Drug Resistance Loci in Plasmodium falciparum

Parasite samples with P. falciparum confirmed by PCR following the methodology previously published [35 (link)] were selected for sequencing of antimalarial drug resistance loci including: pfmdr1 (PF3D7_0523000; including codons 86, 184 and 1246) and pfk13 (PF3D7_1343700; including codons from 412 to codon 723). Genes were PCR amplified following the methodology previously published [34 (link)] with modifications (primers and thermocycling conditions in Table S1) and products cleaned using SureClean (Bioline, USA) following manufacturer’s instructions. The PCR products were analyzed by electrophoresis on a 2% agarose gel stained with GreenSafe (Nzytech, Portugal) to confirm amplification. The PCR products were sequenced using Sanger capillary platform at Eurofins Genomics, Germany, and the resulting sequences were analyzed using BLAST: Basic Local Alignment Search Tool (https://blast.ncbi.nlm.nih.gov/Blast.cgi; accessed on 18 August 2021). The laboratory-adapted 3D7 clone (MRA-102) was used as reference.