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. 2018 Jan 10;8:270. doi: 10.1038/s41598-017-18442-3

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© The Author(s) 2017

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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Parameters for transfection of Trichomonas vaginalis for detection of nanoluciferase activity. (A) Cartoon representation of nanoluciferase plasmids and predicted outcome for protein production. (B) A total of 1 × 10⁷ parasites were either nucleofected using the U-033 (“U”) or D-023 (“D”) programs (Amaxa) or mock (no nucleofection) with 10 μg of pMN::nluc or pMN::nluc(stop) in buffers Parasite-1 or T-cell (Lonza) or electroporated (975 μF, 350 V, Bio-Rad) in completed TYM media. Plasmids are pMN::nluc, denoted as “nluc” and pMN::nluc(stop), denoted as “nluc(stop)” where indicated. Parasites were immediately recovered in completed TYM media. After 24 hours, a total of 1 × 10⁶ parasites were collected and nanoluciferase activity was measured by NanoGlo assay (Promega). Graph represents the average relative luminescence value +/− standard deviation of two transfections per condition. (C) Samples transfected with the pMN::nluc or pMN::nluc(stop) constructs and the U-033 program from Fig. 2B were subjected to G418-selection and allowed to grow for an additional +24 hours (+48 hours total, post-transfection). Equivalent numbers of parasites (1 × 10⁶) were assayed for nanoluciferase activity and the graph represents the average relative luminescence value +/− standard deviation of two attempts per condition.