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. 2020 Jun 16;10:9706. doi: 10.1038/s41598-020-66537-1

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© This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply 2020

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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A parallel SELEX scheme was employed to select aptamers to P. falciparum-infected red blood cells (RBCs). (i) Combinatorial ssDNA libraries, consisting of oligonucleotides with a centrally randomized region of 45 nucleotides flanked by specific primers, are removed of sequences that bind to uninfected RBCs in a negative selection step. (ii) The depleted libraries are incubated with P. falciparum-infected RBCs to enforce selection of aptamers binding determinants unique to infected erythrocytes (IEs). (iii) Following thorough washing, bound sequences are amplified by PCR. (iv) Aptamers are regenerated as ssDNA either by alkaline denaturation or digestion with lambda exonuclease. The resulting enriched ssDNA pools are used in the next round of the negative selection, positive selection, and amplification. In the 10^th SELEX round, the selected ssDNA from the two independent selections are analyzed for aptamer enrichment using the Illumina high throughput sequence platform.