Supplementary Figure 3: Expression of TALE against telomeres had no effect on cell-cycle profile or karyotype.

(a) Two ES cell-lines stably expressing TALE-mClover_telomere (E14Telo1 and E14Telo2), TALE-mRuby_MajSat, and H2B-tdiRFP, and perental E14 were analyzed by FACS. Histograms of PI staining are shown. (b) FACS data was analyzed using ModFit to classify cell-cycle phases into S, G2/M, and G1 phase. The proportion of each phase for the indicated cell-lines is shown. Note that the proportion of cells in each cell-cycle is indistinguishable between parental E14 and the two clones expressing TALE-mClover_telomere. Shown is a representative experiment of two independent biological replicates. (c) Percentage of cells displaying normal karyotype consisting of 40 chromosomes. More than 80% of the cells from the clones stably expressing TALE-mClover_telomere displayed normal karyotype, similarly to the parental ES cell line, indicating that chromosome segregation was not affected by expression of the TALE-mClover_telomere. n indicates the number of cells analyzed for each clone. (d and e) Relative telomere length is analyzed by Q-FISH. (d) Representative images of indicated cell-line showing fluorescent signal for telomere Q-FISH (magenta) and DAPI (gray). (e) Relative telomere length is shown with s.d. n indicates number of chromosome spread analyzed. *, P = 0.047, ** P = 0.087 (Student's t-test). (f) Quantitative RT-PCR analysis for TALE-mClover_telomere in indicated cell-lines. PCR was normalized using Gapdh mRNA levels. Shown are the mean ± s.d., n = 3.