Integrative transcriptome sequencing identifies trans-splicing events with important roles in human embryonic stem cell pluripotency

(Downloading may take up to 30 seconds. If the slide opens in your browser, select File -> Save As to save it.)

Click on image to view larger version.

Figure 3.
Figure 3.

Effect of tsRMST knockdown on pluripotency maintenance of hESCs. (A) Alkaline phosphatase staining and quantification of hESCs transfected with control (shLuc) or shTS2 virus at 4 and 7 d post-viral transduction. (Scale bar) 200 μm. (B) Heat map clustering analysis of genes related to pluripotency, neuroectoderm, mesoderm, and endoderm in shLuc- and shTS2-transduced hESCs. Relative fold changes are listed. Green and red values represent fold change for down- and up-regulation, respectively. (C) qRT-PCR analysis of RNA isolated from hESCs transfected with control or shTS2 virus at 4 and 7 d post-viral transduction, to detect pluripotency-related genes (NANOG, POU5F1, SOX2, TCF7L1), and lineage-specific genes, including mesodermal markers (T, GSC, MIXL1), endodermal markers (SOX17, GATA4, GATA6, SOX7), and neuroectodermal markers (PAX6, SOX1). (D) Fluorescence-activated cell sorting (FACS) analysis of NANOG+ and POU5F1+ cell populations in hESCs transfected with shLuc or shTS2 virus at 4 and 7 d post-viral transduction. Three independent transfections were performed to determine the mean. (E) Immunocytochemistry analysis and quantification of the expression of NANOG, POU5F1 (pluripotency markers), T (mesodermal marker), SOX17 (endodermal marker), and PAX6 (ectodermal marker) in shTS2-transduced hESCs at 4 and 7 d post-viral transduction. (Scale bar) 20 μm. (F) FACS analysis of NANOG+ and POU5F1+ cell populations in shTS2 virus-transfected hESCs and shTS2-rescue hESCs. Three independent transfections were performed to determine the mean. (G) Immunocytochemistry analysis and quantification of the expression of NANOG and POU5F1 (pluripotency markers) in shTS2 virus-transfected hESCs and shTS2-rescue hESCs. (Scale bar) 20 μm. (H) qRT-PCR analysis of pluripotency-related genes (POU5F1, NAONG, SOX2, TCF7L1) in RNA isolated from shTS2 virus-transfected and shTS2-rescue hESCs. (I) Alkaline phosphatase staining and quantification of shTS2 virus-transfected hESCs with or without tsRMST coexpression (shTS2-rescue). (Scale bar) 200 μm. All indicated P-values were estimated by the two-tailed two-sample t-test. Significance: (*) P < 0.05; (**) P < 0.01; and (***) P < 0.001.

This Article

  1. Published in Advance October 16, 2013, doi: 10.1101/gr.159483.113 Genome Res. 24: 25-36

Preprint Server



Current Issue

  1. January 2026, 36 (1)
  1. Current Issue
  1. Alert me to new issues of Genome Research