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. 2019 Apr 23;9:6467. doi: 10.1038/s41598-019-42914-3

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© The Author(s) 2019

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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Differential activity of RRE and Rev sequences. (a) Three different 234-nt RREs were cloned into the eGFP/TagBFP HIV-derived construct, including RREs from NL4-3 and two derived from patient isolates (SC3-M0A and SC3-M57A)²³. 1000 ng of the HIV-derived construct was transfected into 8 × 10⁵ 293T/17 cells along with 0, 25, 50, or 100 ng of pCMV-SC3-(M0-B/M57-A) Rev²³. Twenty-four hours after transfection, flow cytometry was performed and the ratio of eGFP/TagBFP signal was determined as a measure of Rev-RRE functional activity. The maximum measured activity was set as 100. (b) Three Rev sequences from NL4-3 or two different primary isolates^53,54 were cloned into the MSCV vector carrying an mCherry fluorescent marker. 1000 ng of an eGFP/TagBFP HIV-derived construct containing an NL4-3 RRE sequence was transfected into 8 × 10⁵ 293T/17 cells along with 12.5, 25, 50, 100, or 200 ng of a pMSCV-Rev-IRES-mCherry construct. Flow cytometry was performed 24 hours after transfection and the data were analyzed as above. N = 2, bars, which are too small to be seen in this figure, represent SEM for both graphs.