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. 2017 May 8;7:1550. doi: 10.1038/s41598-017-01740-1

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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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MB-based imaging of single HOTAIR transcripts in the nucleus and the cytoplasm. (a) Images of anti-repeat MBs for the detection of single pHOTAIR-8x transcripts. Following microporation of NIH3T3 cells with or without pHOTAIR-8x with 5 µM of anti-repeat MBs, the cells were fixed and permeabilized and smFISH was performed to assess the accuracy of MBs for detecting single RNA transcripts. Representative maximum intensity projection images of MB and smFISH signals are shown. Note that in NIH3T3 cells not transfected with pHOTAIR-8x, lack of smFISH signal shows the specificity of the smFISH probes for the human HOTAIR transcript used. (Scale bar, 10 µm) (b) Diffusion kinetics of single pHOTAIR-8x lncRNA transcripts in HeLa cells. HeLa-HOTAIR-8x cells were microporated with 5 µM anti-repeat MBs, and time-lapse images were acquired 8 h after microporation. Single particle tracking analysis was performed to determine the diffusion coefficient of individual RNAs as described in Materials and Methods. The distribution of diffusion coefficients of single mobile pHOTAIR-8x RNAs in the nucleus (n = 113 tracks) and in the cytoplasm (n = 255 tracks) analyzed from 30 cells. Inset shows the mean ± SE diffusion coefficients.