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. Author manuscript; available in PMC: 2016 May 12.
Published in final edited form as: Nat Biotechnol. 2016 Apr 18;34(5):528–530. doi: 10.1038/nbt.3526

Figure 2.

Figure 2

Localization and live tracking of multiple chromosome-specific loci simultaneously. (a) Construct pCRISPRainbow-sgRNA-Cx-C14-C7-C1-C13-C3 for co-expression of six sgRNAs. sgRNA-2XMS2-Cx, sgRNA-2XPP7-C14, sgRNA-2XboxB-C7, sgRNA-MS2-PP7-C1, sgRNA-PP7-boxB-C3, and sgRNA-boxB-MS2-C3 were constructed individually and then subcloned into a single vector. (b) pCRISPRainbow-sgRNA-Cx-C14-C7-C1-C13-C3, dCas9, MCP-3XBFP, PCP-3XmNeonGreen, and N22-3XRFP were co-transfected into U2OS cells. Each CRISPRainbow color was dedicated to one chromosome locus: blue for chromosome X (Cx), green for chromosome 14 (C14), red for chromosome 7 (C7), cyan for chromosome 1 (C1), yellow for chromosome 13 (C13) and magenta for chromosome 3 (C3), respectively. Scale bar, 3 μm. (c) Unique sites in chromosome 3, 9, and 13 as well as telomeres were labeled simultaneously using the CRISPRainbow colors blue, green, red, and yellow, respectively. The movements of these loci were recorded at 50 ms per frame for 10 s (200 total frames). All trajectories were shifted to start from the origin (0, 0) for easy comparison of the movement vectors. Scale bar, 5 μm. Data in all panels are representative of experiments performed at least three times.